Open access

Removal of Acrylamide by Microorganisms

Written By

Jittima Charoenpanich

Submitted: July 4th, 2012 Published: October 2nd, 2013

DOI: 10.5772/56150

Chapter metrics overview

3,428 Chapter Downloads

View Full Metrics

1. Introduction

Acrylamide (CH2=CHCONH2) is a well-known bifunctional monomer, appearing as a white odorless flake-like crystal. It is soluble in water, methanol, ethanol, dimethyl ether, and acetone, but insoluble in benzene and heptane. Acrylamide is incompatible with acids, bases, oxidizing agents, irons and iron salts. It decomposes non-thermally to form ammonia while thermal decomposition produces carbon monoxide, carbon dioxide, and oxides of nitrogen [1].

As a commercial conjugated reactive molecule, acrylamide has been used worldwide for the synthesis of polyacrylamide and other polymers [2, 3]. It has also been used as a binding, thickening, or flocculating agent in grout, cement, sewage, wastewater treatment, pesticide formulation, cosmetics, sugar manufacturing, and to prevent soil erosion. Polymers of this compound have been used in ore processing, food packaging, plastic products, and in scientific and medical laboratories as solid support for the separation of proteins by electrophoresis [4]. Acrylamide monomer is also widely used as an alkylating agent for the selective modification of sulfhydryl proteins and in fluorescence studies of tryptophan residues in proteins. In 2002, there was an alarming report of the occurrence of acrylamide at high levels up to 3 mg/kg in plant-derived foods and thought to form during cooking allowing the formation of Maillard browning products [5]. Many reports have suggested that acrylamide seems to be found in foods that have been processed by heat-treatment methods other than boiling [6]. One possible pathway to the formation of acrylamide is via the Maillard reaction between amino acids, particularly asparagines, and reducing sugars at high temperatures [5, 6]. Some reports suggest acrylamide could form by acrolein (2-propenal, CH=CHCHO), a three-carbon aldehyde, by either the transformation of lipids or the degradation of amino acids, proteins and carbohydrates [7-12].

Acrylamide could be absorbed through unbroken skin, mucous membranes, lungs, and the gastrointestinal tract. Human exposure to acrylamide is primarily occupational from dermal contact with the solid monomer and inhalation of dust and vapor. Although it is not toxic in polymer form, the monomer can cause peripheral neuropathy. Residual monomer in polymers is also of health concern [13]. Primary exposure occurs during the handling of monomers. Two acrylamide manufacturing factories showed breathing zone concentrations of 0.1 to 3.6 mg/m3 [1]. During normal operations, workers at another plant were exposed to not more than 0.3 mg/m3. Aside from occupational exposure, probable exposure to the general public is through consumption of certain foods [14]. Another source of acrylamide exposure to the general public could be through drinking water treated with polyacrylamide flocculants [13]. Acrylamide may not be completely removed in many water treatment processes with some remaining after flocculation with polyacrylamides probably due to its water solubility and is not absorbed by sediment [15].

Acrylamide is evidentially a neurogenic, terratogenic or carcinogenic toxicant in animals [16]. The neurotoxic properties of acrylamide have been studied for humans in relation to occupational exposures and, experimentally, in laboratory animals. Understanding of acrylamide-induced neuropathies is quite advanced, a consequence of more than 30 years of research on the possible mechanisms of action [17]. The mechanism underlying the neurotoxic effects of acrylamide as with other toxins are interference with the kinesin-related motor proteins in nerve cells or with fusion proteins in the formation of vesicles at the nerve terminus and eventual cell death [18]. Neurotoxicity and resulting behavioral changes in acrylamide-exposed laboratory animals can reduce reproductive fitness. Further, kinesin motor proteins are important in sperm motility, which could alter reproductive parameters. Effects on kinesin proteins could also explain some of the genotoxic effects on acrylamide. These proteins form the spindle fibers in the nucleus that function in the separation of chromosomes during cell division. This could explain the clastogenic effects of the chemical noted in a number of tests for genotoxicity and assays for germ cell damage [4].

Advertisement

2. Release of acrylamide in environment

Acrylamide is a synthetic monomer with a broad spectrum of industrial applications, mainly as a precursor in the production of several polymers, such as polyacrylamide [1, 19]. High molecular weight polymers can be modified to develop nonionic, anionic, or cationic properties for specific uses [1, 20]. Various grades of acrylamide are available with the industrial grade typically with a purity of 98 to 99%. Acrylamide for laboratory use ranges from routine to pure, the former for electrophoresis, the latter for molecular applications [21]. The largest demand for acrylamide polymers in industry is for flocculation of unwanted chemical substances in water arising from mining activities, pulp and paper processing, sewage treatment, and other industrial processes. Applications are based on the principles of colloidal suspensions and used to clean up liquids, particularly aqueous media, either for disposal or human consumption [20, 22-23]. Acrylamide is also used as a chemical intermediate in the production of N-methylol acrylamide and N-butoxy acrylamide and as a superabsorbent in disposable diapers, medical, and agricultural products [24]. Small amounts of acrylamide are also used in sugar beet juice clarification, adhesives, binders for seed coatings and foundry sand, printing ink emulsion stabilizers, thickening agents for agricultural sprays, latex dispersions, textile printing paste, and water retention aids [25]. An aqueous 50% solution of acrylamide is used as acrylic copolymer dispersions in surface coatings and adhesives. In surface coatings, polymers are used as dispersants and binders to provide better pigment separation and flow. Surface coatings are used on home appliances and in the automotive trade [1, 26]. In addition, polyacrylamide has been used in both paper production process and treatment of mill wastewater [27]. Emulsions of polyacrylamide, calcium carbonate and clay are applied as a white coating in the manufacture of cardboard cartons [22]. These polymers are used as thickeners in soap and cosmetic preparations, and in skin care and hair grooming products, to impart a smooth after-feel and shine [22]. For oil drilling, liquid or powder partially-hydrolyzed polyacrylamide is used as additives to water based drilling mud to provide a lubricating film and reduce friction at the drill bit, impart stability to shale and clay and increase viscosity [1, 22, 26]. Moreover, specialized gels comprised in part of acrylamide polymers are manufactured for use as lubricants in the textile dying components to which fabric or finished garments are added. The gel lubricates the cloth preventing it from clumping together and aids pigment dispersion during the dying process to ensure an even color [22, 28]. In leather processing, acrylamide is used as polymers impart a gloss or specific feel and suppleness to leather. The hide is most commonly placed in a drum with the polymer and various other constituents such as dyes, formaldehyde and pigments, then rolled for about two hours. The polymer can also be applied by brush or spray. There is no set formulation for the components of the mix and the proportion of acrylamide polymer is at the discretion of the operator seeking to obtain the properties required in the tanned product [29-30]. Another major application of acrylamide is to reduce herbicide drift during spray applications. The polymer increases the viscosity of the herbicide solution, allowing for more uniform spray applications, and also increases plant contact time [31-33].

In worldwide usage, acrylamide is released into environment as waste during its production and in the manufacture of polyacrylamides and other polymers. Residual acrylamide concentrations in 32 polyacrylamide flocculants approved for water treatment plants ranged from 0.5 to 600 ppm [13]. Acrylamide may remain in water after treatment [15] and after flocculation with polyacrylamides due to its high solubility and is not readily adsorbed by sediment [34]. Other sources of release to water are from acrylamide-based sewer grouting and recycling of wastepaper. Another important source of contamination is from acrylonitrile-acrylamide production which releases approximately 1 g acrylamide in each liter of effluent [35]. Some reports have indicated that polyacrylamide, in the presence of sunlight and glyphosate, photolytically degrades to acrylamide monomer and this is a direct introduction of acrylamide into agricultural areas [36-38]. The half-life of acrylamide monomer in rivers ranges from weeks to months [22]. However, one report indicates that polyacrylamide does not degrade to acrylamide monomer in the presence of sunlight and glyphosate. Additionally, glyphosate appears to interact with either the acrylamide monomer or polymer, decreasing the rate of monomer degradation [39]. The most important environmental contamination results from acrylamide use in soil grouting [13]. Half-life of acrylamide in aerobic soil increases with decreasing temperature [40]. Under aerobic conditions, acrylamide was readily degraded in fresh water by bacteria with a half-life of 55-70 h, after acclimatization for 33-50 h [41]. Acrylamide has been shown to remain slightly longer in estuarine or salt than fresh water [15].

Acrylamide releases to land and water from 1987 to 1993 totaled over 18.16 tons of which about 85 percent was to water, according to Toxic Chemical Release Inventory of the U.S. Environmental Protection Agency (EPA) [40]. These releases were primarily from plastic industries which use acrylamide as a monomer. In 1992, discharges of acrylamide, reported to the Toxic Chemical Release Inventory by certain US industries included 12.71 tons to the atmosphere, 4.54 tones to surface water, 1,906.8 tones to underground injection sites, and 0.44 tones to land [4]. In an EPA study of five industrial sites that produce acrylamide and polyacrylamide, acrylamide (1.5 ppm) was found in only one sample downstream from a polyacrylamide producer and no acrylamide was detected in soil or air samples [13]. Concentrations of 0.3 ppb to 5 ppm acrylamide have been detected in terrestrial and aquatic ecosystems near industrial areas that use acrylamide and/or polyacrylamides [42-43]. Cases of human poisoning have been documented from water contaminated with acrylamide from sewer grouting. The acrylamide monomer was found to remain stable for more than 2 months in tap water [22]. Atmospheric levels around six US plants were found on an average of < 0.2 µg/m3 (0.007 ppb) in either vapor or particulate form [15]. The vapor phase chemical should react with photochemically produced hydroxyl radicals (half-life 6.6 h) and be washed out by rain [15].

Advertisement

3. Microbial degradation of acrylamide

The interest in environmental problems is continuously growing and there are increasing demands to seek the sustainable and controllable process which do not burden the environment significantly. Biodegradation is one of the classic methods for removal of undesired organic compounds to concentrations that are undetectable or below limits established as acceptable by regulatory agencies.

Acrylamide is likely to partially biodegrade in water within approximately 8-12 days [13]. If released on land, acrylamide can be expected to leach readily into the ground and biodegrade within a few weeks. In five surface soils that were moistened to field capacity, 74-94% degradation occurred in 14 days in three soils and 79 to 80% in 6 days in the other two soils [44]. Acrylamide may not be completely degraded in domestic sewage and water treatment facilities if residence times are relatively short [13, 45]. Further degradation through bioremediation of acrylamide to less harmful substances would alleviate environmental concerns.

Since 1982, microbial degradation of acrylamide has been explored extensively with a diversity of isolates (Table 1), mainly Bacillus, Pseudomonas and Rhodococcus [3, 46-55]. Further, numerous other microorganisms including the representatives of Arthrobacter, Xanthomonas, Rhodopseudomonas, Rastonia, Geobacillus, and a newly family of Enterobacteriaceae [49, 56-62]. Aspergillus oryzae, a filamentous fungal has also been documented as an acrylamide degrader [63].

Several acrylamide degraders use a coupling reaction of nitrile hydratase (EC 4.2.1.84) and amidase (EC 3.5.1.4) for biotransformation of acrylonitrile to acrylic acid via acrylamide as an intermediate [46, 56]. For example, R. rhodochrous J1 changed acrylonitrile to acrylamide and subsequently to acrylic acid [47] and R. erythropolis utilized either 2-arylpropionamides or acrylamide to form acrylic acid and ammonia [64]. In China, Nocardia sp. 163, a soil derived bacterium from Taishan Mountain harboring the highest nitrile hydratase activity on acrylonitrile was also used frequently for bioconversion of acrylamide [65]. Another prominent example is Rhodococcus sp. AJ270 which is a powerful and robust nitrile hydratase/amidase-containing microorganism isolated by Guo et al [66]. An aliphatic amidase (amidohydrolase) has been found to be the responsive enzyme for the deamidation of acrylamide to acrylic acid and ammonia [50, 59, 62, 64-67].

In 1990, Shanker and his colleagues isolated an acrylamide-degrading bacterium, Pseudomonas sp., from soil using an enrichment method. This bacterium degraded high concentration of acrylamide (4 g/l) to acrylic acid and ammonia. An amidase was also found to be the relevant enzyme for the hydrolysis of acrylamide and other short chain aliphatic amides like formamide and acetamide but not on acrylamide analogues, methacrylamide and N, N-methylene bisacrylamide [48].

Many aerobic microorganisms utilize acrylamide as their sole source of carbon and energy including Pseudomonas sp. and Xanthomonas maltophilia. Nawaz and his team found amidase in cell free extracts of these species and suggested it was involved in acrylamide degradation [49]. This is consistent with their earlier conclusion of acrylamide degradation by Rhodococcus sp. [50]. Later, the denitrifying bacteria, Pseudomonas stutzeri was found to use acrylamide as substrate in the acrylonitrile–butadiene–styrene resin wastewater treatment system. The strain could remove acrylamide at concentrations below 440 mg/l under aerobic conditions [52]. Acclimation of microorganisms is believed to enhance acrylamide biodegradation. Complete degradation of acrylamide at 10–20 ppm in river water occurred in about 12 days with non-acclimated microorganisms, but in only 2 days with acclimation [3]. In 2009, scientists in Malaysia reported two acrylamide-degrading bacteria, Bacillus cereus DRY135 and Pseudomonas sp. DRYJ7. Acrylic acid was also detected as a metabolite in the degradation [53-54]. Aspergillus oryzae KBN 1010 has been the only fungi documented as an acrylamide degrader [63].

In domestic wastewater in Thailand, four novel acrylamide-degrading bacteria (Enterobacter aerogenes, Kluyvera georgiana, Klebsiella pneumoniae, and Enterococcus faecalis) were isolated. E. aerogenes and K. georgiana showed degradation potential of acrylamide up to 5000 ppm at the mesophilic temperatures and could degrade other aliphatic amides especially short to medium-chain length but not amide derivatives [60-61]. Removal of acrylamide and ammonium nitrogen from industrial wastewater by E. aerogenes was generally higher than that by mixed cultures of microorganisms [68].

Microorganisms Source Conditions Reference
Bacteria
Pseudomonas chlororaphis B23 Soil Aerobic (Enzymatic degradation) [46]
Arthrobacter sp. J-1 Soil Aerobic (Enzymatic degradation) [56]
Rhodococcus rhodochrous J1 Soil Aerobic (Free cells) [47]
Pseudomonas sp. Soil Aerobic (Free cells) [48]
Pseudomonas sp.
Xanthomonas maltophilia
Soil Aerobic (Immobilized cells) [49]
Rhodococcus sp. Soil Aerobic (Enzymatic degradation) [50]
Rhodococcus erythropolis MP50 Soil Aerobic (Enzymatic degradation) [64]
Rhodococcus sp. Soil Aerobic (Immobilized cells) [51]
Pseudomonas stutzeri Wastewater treatment system Aerobic (Free cells) [52]
Rhodopseudomonas palustris Bovine slaughterhouse Photoheterotropic (Free cells) [57]
Pseudomonas aeruginosa Soil Aerobic (Free and immobilized cells) [3]
Ralstonia eutropha TDM-3 Wastewater treatment system Anaerobic (Free cells) [58]
Bacillus cereus DRY135 Soil Aerobic (Free cells) [53]
Pseudomonas sp. DRYJ7 Antarctic soil Aerobic (Free cells) [54]
Natural microbial populations Rocky Ford Highline Canal, Colorado USA Aerobic and anaerobic (Free cells) [69]
Ralstonia eutropha AUM-01 Soil Aerobic (Free cells) [59]
Enterobacter aerogenes Domestic wastewater Aerobic (Free and immobilized cells) [60]
Kluyvera georgianaKlebsiella pneumoniaeEnterococcus faecalis Domestic wastewater Aerobic (Free cells) [61]
Geobacillus thermoglucosidasius AUT-01 Soil Aerobic (Free cells) [62]
Pseudomonas aeruginosa DS-4 Soil Aerobic (Free cells) [55]
Fungi
Aspergillus oryzae KBN 1010 Filamentous fungi used in food and beverage industries Aerobic (Free cells) [64]

Table 1.

Acrylamide-degrading microorganisms.

Degradation of acrylamide under anaerobic conditions has been rarely described. Recently a new strain of Rhodopseudomonas palustris was found capable of using acrylamide under photoheterotrophic conditions but grew poorly under anaerobic dark or aerobic conditions. A study of acrylamide metabolism by nuclear magnetic resonance showed the rapid deamidation of acrylamide to acrylate and further to propionate [57]. More recently, the denitrifying bacterium, Ralstonia eutropha TDM-3 isolated from the wastewater treatment system associated with the manufacture of polyacrylonitrile fiber consumed acrylamide to concentration of 1446 mg/l, above which it was toxic [58]. This report is similar with the potential of soil bacteria, Ralstonia eutropha AUM-01 and Geobacillus thermoglucosidasius AUT-01 [59, 62]. One report, and perhaps most interesting, removal of acrylamide has been found potentially with the natural microbial populations in Rocky Ford Highline Canal, Colorado USA [69]. Degradation of acrylamide occurs under aerobic or anaerobic conditions, with nitrate serving as the most favorable anaerobic electron acceptor. Phylogenetic analysis of these cosmopolitan microorganisms suggest the potential for biodegradation in similar lotic systems such as Pseudomonas, Rhodococcus, and Bacillus. New proteobacterial genera (Pectobacterium, Citrobacter, Delftia, Comomonas, and Methylobacterium) were also found [69]. Microbial degradation of a lipid in conjunction with acrylamide was also report with Pseudomonas aeruginosa DS-4. Salad oil was believed to be an essential factor for acrylamide biodegradation by this bacterium. The degradation rate of acrylamide was affected by the incubation time of the acclimated strain DS-4. Longer incubation time with acrylamide resulted in more efficient degradation [55].

Advertisement

4. Metabolism of acrylamide

Until now, we can not deny possible routes for acrylamide other than deamination via amidase [50, 59, 62, 64, 67]. The subsequent fate of acrylate is not well understood but probably involves pathways and enzymes that have been characterized to various degrees for other acrylate utilizing bacteria (Figure 1). Acrylate metabolism is believed to proceed via hydroxylation to β-hydroxypropionate, then oxidized to CO2 [48] or reduced to propionate [57]. Another plausible pathway for mineralization of acrylamide is via formation of acrylyl CoA which eliminates lactate as a final product [48].

A powerful tool that also enables unraveling acrylamide metabolic pathways is the sequential induction of catabolic enzymes and intermediatary metabolites. Further, insight into degradative pathways is also provided from assaying the probable key proteins that are synthesized at sufficient levels when acrylamide is present. Using proteome analysis, fifteen proteins differentially expressed from Enterobacter aerogenes grown on acrylamide were identified. Six protein homologues with amidohydrolase, urease accessory protein, quaternary ammonium compound resistance proteins, dipeptide transport protein, Omp36 osmoporin and large conductance mechanosensitive channel proteins (MscL) are seemingly involved in acrylamide stress response and its degradation. Five proteins identified as GroEL-like chaperonin, ArsR-transcriptional regulator, Ts- and Tu-elongation factor and trigger factor and four proteins (phosphoglycerate kinase, ATP synthase β-subunit, malate dehydrogenase and succinyl-CoA synthetase α-subunit) are expected to be relevant to adaption of E. aerogenes in the presence of acrylamide [70]. Based on the results, Charoenpanich and Tani have proposed acrylamide may be assimilated using Omp36 osmoporin and dipeptide transport proteins. Acrylamide is toxic, indeed lethal, to most microorganisms, however some bacteria have adapted their metabolism to use this substance as an energy source. Important to this adaptation is the evolution of genes that encode amidohydrolase (amidase) and other synthesis proteins that deaminate acrylamide to acrylic acid and ammonium [48, 50-51, 60-61]. With this, acrylic acid can be changed to propionate and subsequently succinyl CoA [57, 71-72] to generate energy. Potentially harmful ammonium is detoxified and with MscL protein and released from the cell [70].

Figure 1.

Possible biological fates of acrylate produced from acrylamide deamidation.

Advertisement

5. Bioremediation of acrylamide and future prospects

Bioremediation is viewed as a sustainable process for wastewater treatment, which under appropriate conditions, can promote an efficient reduction of organic matter with minimal energy requirements and, therefore, low costs. Major limitations are the bioavailability of the organic matter and the finding of efficient biodegraders. Physico-chemical environmental conditions also greatly influence the rate and extent of degradation. In general, degradation efficiency is dependent on three overall factors (i) microorganisms that can degrade the specific chemical structure (ii) environmental conditions that allow the microorganisms to grow and express their degradation enzymes and (iii) good physical contact between the organic substrate and the organism.

Rapid degradation of acrylamide coupled with growth requires not only amidase or microorganism producing amidase, but also a whole pathway, i.e. a set of enzymes that are differentially synthesized in the presence of acrylamide. Although a complete catabolic pathway for acrylamide does not exist, recombination and mutation processes and exchange of genetic information between microorganisms may lead to the development of organisms with improved catabolic activities. Alternatively, microorganisms can cooperate by combining their catabolic potential in mixed cultures and in this way may completely mineralize acrylamide. Wang and Lee elucidated the effectiveness of Ralstonia eutropha TDM-3 and mixed cultures of wastewater from the manufacture of polyacrylonitrile fiber in treating acrylamide in synthetic wastewater. They found that mixed culture and R. eutropha TDM-3 can jointly consume acrylamide up to concentrations of 1446 mg/l and completely remove acrylamide with a sufficient supply of nitrate as electron acceptors [58]. A similar result has been found in E. aerogenes. If grown with mixed cultures from a municipal wastewater treatment plant, they can completely and rapidly convert acrylamide to acrylic acid [68]. Acrylamide up to 100 mg/L can efficiently be removed from amended canal water and sediment slurries under aerobic conditions. Using natural nitrate-reducing microorganisms in a canal environment, potential fate of acrylamide (70.3-85%) was found after 60 days [69].

Microorganisms typically require sufficient water, inorganic nutrients, carbon sources, and trace elements for maintenance and growth. Besides growth substrates, other specific organic compounds such as vitamins or other growth factors are essential for some microorganisms. Monosaccharides like glucose and fructose have been reported as support elements for the growth and degradation potential of acrylamide-degrading bacteria [53-54]. However, in some cases supplementation of acrylamide containing growth medium with glucose or succinate as additional carbon source demonstrated a severe repression in degrading ability [48, 71-75]. Addition of glutamate or ammonium sulfate as an additional nitrogen source to the growth medium demonstrated an increase in degradation potential compared to the cells grown only on acrylamide [48]. One interesting study found that Pseudomonas aeruginosa DS-4 isolated from lipid wastewater required salad oil for growth and acrylamide degradation [55].

Toxic compounds (e.g. heavy metals) should not be present at high concentrations, since they can inactivate essential enzymes. As explained in [51] iron (<10 mM) enhanced the rates of acrylamide degradation of Rhodococcus sp. but copper, cobalt and nickel inhibited the degradation. Mercury and chromium inhibited acrylamide degradation by Pseudomonas aeruginosa while nickel at lower concentrations (200 and 400 ppm) improved the degrading ability [3].

Optimum conditions for acrylamide biodegradation are achieved if pH and temperature are in the range of pH 6-8 and mesophilic temperature (15-30ºC), respectively [3, 45-48, 53-55]. Most microorganisms consume considerably less energy for the maintenance of basic functions under neutral conditions. This means that more energy is available for growth. It has been known that metabolic activity of tropical soils typically is high and fosters several processes such as carbohydrate fermentation and carbon dioxide production leading to the lowering of pH. Thus, for successful bioremediation of pollutants including acrylamide pH control may be essential. Addition of an inexpensive chemical such as calcium carbonate to neutralize soil pH during bioremediation can optimize remediation [76].

Studies on acrylamide biodegradation are mainly concerned with the isolation and identification of suitable microbial strains. Most studies use either free or immobilized cells for acrylamide removal. Of these, immobilized cells are advantageous because the immobilized cells are less likely than free cells to be adversely affected by predators, toxin, or parasites [77-78]. Additionally, they can be reused, saving resources and time. However, the implementation of immobilized cells may be sensitive to pH, temperature and acrylamide concentration. Moreover, large accumulations of the metabolic intermediate, acrylic acid, may affect some microbial activity [3, 51, 60]. Hence, the attempt to biotransform acrylamide with amidase or nitrile-converting enzymes via hydrolysis.

Microbial degradation of nitriles proceeds through two enzymatic pathways. Nitrilase (EC 3.5.5.1) catalyzes the direct cleavage of nitriles to the corresponding acids and ammonia, and nitrile hydratase (NHase) catalyzes the hydration of nitriles to amides. Both nitrile-converting enzymes have increasingly attracted attention as catalysts for processing many organic chemicals [79-81]. Nitrile hydratase is commonly used as the catalyst in the production of acrylamide and is known as one of the most important industrial enzymes [82-83]. Generally, the gene operon of nitrile hydratase consists of the genes for the alpha and beta subunits of NHase, the NHase activator and amidase. The amides produced by NHase are degraded to their corresponding free carboxylic acids and ammonia by the action of amidases [84]. Thus, nitrile-converting enzymes are of broad use as alternatives for acrylamide biotransformation.

Acrylic acid, the intermediate product in acrylamide catabolism, is a commodity chemical with an estimated annual production capacity of 4.2 million metric tons [85]. Acrylic acid and its esters can be used in paints, coatings, polymeric flocculants, paper and so on. It is conventionally produced from petrochemicals. Currently, most commercial acrylic acid is produced by partial oxidation of propene which produces undesirable by-products and large amount of inorganic wastes [86]. Currently, there is an innovative manufacturing method using nitrile-amide converting enzymes. For acrylamide degraders, it is initially degraded to ammonia and acrylic acid (acrylate), a process catalyzed by amidase. Then acrylate is reduced to generate energy for growth. Until now, the acrylate-utilizing enzyme has not been well characterized but believed to be acrylate reductase [48, 57]. The identification of the gene encoding this enzyme remains a challenge. Moreover, from an economic aspect, the acrylate reductase-deficient strains created by a gene-disruption method, lead to acrylic acid accumulation in wastewater and are recommended for acrylamide bioremediation in the future.

Sequence similarities have been identified using computer methods for database searches and multiple alignment, between several nitrilases, cyanide hydratase, β-alanine synthase and the first type of aliphatic amidases which hydrolyze only short-chain aliphatic amides [87]. All these enzymes involving the reduction of organic nitrogen compounds and ammonia production exhibited several conserved motifs. One of which contains an invariant cysteine that is part of the catalytic site in nitrilases. Another highly conserved motif includes an invariant glutamic acid that might also be involved in catalysis. Sequence conservation over the entire length of these enzymes, as well as the similarity in the reactions constitutes a definite family which points to a common catalytic mechanism [88]. Chemical mutagenesis and X-ray crystallography have been analyzed for three-dimensional structures of amidases. Only a few crystal structures of nitrilase-related amidases have been reported with Pseudomonas aeruginosa amidase the first [89-90]. The three dimensional-structures showed a conserved α-β-β-α sandwich fold resembling the conserved structural fold of the nitrilase superfamily structures. Analysis of the three dimension-structures identified E59, K134, and C166 as a catalytic triad [89]. Similar catalytic triad residues were also reported in the three dimensional structural models of amidase from Rhodococcus erythropolis, Helicobacter pylori, and Bacillus stearothermophilus [89] and also in the amidase of novel acrylamide-degrading Enterobacter aerogenes [91]. The crystal structure of Xanthomonas campestris XC1258 amidase showed a monomeric structure of globular α/β protein comprising mainly six α helices and two six-stranded β-sheet (Figure 2). This is the typical nitrilase-superfamily α-β-β-α fold. The hexamer preserving the eight-layered α-β-β-αα-β-β-α structure in holoenzyme across an interface has also been reported [92]. The analysis of small asymmetric catalytic site of the Geobacilus pallidus RAPc8 amidase suggested that access of a water molecule to the catalytic triad (C, E, K) side chains would be impeded by the formation of the acyl intermediate. The conserved E142 in the catalytic site acts as a general base to catalyze the hydrolysis of this intermediate [93]. This confirmed the conservation of the E, K, C catalytic triad across the nitrilase superfamily members and also supported the classification of the amidases in the nitrilase superfamily.

Figure 2.

(a) The monomeric tertiary structure of amidase from Xanthomonas campestris XC1258, color-coded from blue (N-terminal) to red (C-terminal), and (b) the primary sequence of XC1258 amidase. Reprinted from Ref. [92].

Acrylamide amidases have similar sequences with nitrilases and seem to have descended from a common ancestry along with members of the sulfhydryl enzyme family. In these amidases an invariant cysteine residue was reported to act as the nucleophile in the catalytic mechanism and is confirmed by the three dimensional structural model of the amidase of Pseudomonas aeruginosa. This was built by comparative modeling using the crystal structure of the worm nitrilase fusion protein, NitFhit as the template. The putative catalytic triad C-E-K is conserved in all members of the nitrilase superfamily [89]. The signature amidases possesses two real active site residues D191 and S195 among the various conserved residues within the signature sequence common to all enantioselective amidases. D191N and S195A substitutions in Rhodococcus amidase has been shown to completely suppress amidase activity [94-95]. These sequences are also present within the active site sequences of aspartic proteinases. Thus, amide bond cleaving enantioselective amidases that are coupled with nitrile hydratases are evolutionary related to aspartic proteinases. Further structural characterization of the amidase produced by acrylamide-degrading bacteria should reveal what other differences are present. It may be possible to use this information to aid protein engineering of the enzymes in order to improve their efficiency and specificity.

Development of thermostable amidase is also important. Based on the three-dimensional structure of amidase, additional disulfide bridges can be engineered by site-directed mutagenesis for enzyme stabilization. Novel amidases that show broad substrate specificity may be developed to biodegrade the toxic environmental pollutants, acrylamide and amides. Random approaches such as directed evolution, reverse engineering and site-directed mutagenesis could be applied to achieve such ends.

Our understanding of the biochemistry and molecular biology of amidase is advancing rapidly and already providing information that is of use today. Moreover, recent developments in amidase studies have broadened the scope of potential applications of the enzyme in acrylamide bioremediation as well as that of acrylic acid production. I predict that these developments combined with progress in genetic engineering and enzyme crystallography will have a major effect on the practical applications of acrylamide bioremediation.

Advertisement

6. Concluding remarks

A huge demand for acrylamide as an ubiquitous monomer for industry led to its environmental presence, however the International Agency for Research on Cancer has classified this compound as a probable human carcinogen. Bioremediation seems to be the only efficient and environmentally friendly process to decompose this monomer. The first step in developing acrylamide bioremediation is to choose high potent microorganisms. Choice of microorganisms is challenging owing to the large scale degradation of acrylamide and elucidation of the intermediate in catabolic pathways is the first important step. Nevertheless, the main problem is the rapid conversion of intermediate acrylic acid to other metabolites. Research on the relationship between degradation mechanisms and membrane structure of acrylamide-utilizing bacteria awaits further characterization. It is noteworthy that successful remediation of acrylamide depends on the ability of microbes to adapt to new environmental conditions and the availability of active and stable chemical degrading bacteria. Indigenous predators, parasites and toxicants are known to severely restrict biodegradation and should be a concern.

Advertisement

Acknowledgments

The author is grateful to Dr. N. Kurukitkoson for his encouragement to write this review and would like to thank F.W.H. Beamish for proofreading the manuscript.

Advertisement

Nomenclature

Amino acids

E: Glutamic acid

K: Lysine

C: Cysteine

D: Aspartic acid

N: Asparagine

S: Serine

A: Alanine

References

  1. 1. IARCSome Chemicals Used in Plastics and Elastomers. In: IARC Monographs on the evaluation of carcinogenic risk of chemicals to human. France: International Agency for Research on Cancer; 1986403
  2. 2. PrasadD. YPolyacrylamide as a coagulant aid in water treatment. Chemical Age of India 198234387
  3. 3. PrabuC. SThatheyusA. JBiodegradation of acrylamide employing free and immobilized cells of Pseudomonas aeruginosa. International Biodeterioration and Biodegradation 20076069
  4. 4. ExonJ. HA review of the toxicology of acrylamide. Journal of Toxicology and Environmental Health 20069397
  5. 5. MottramD. SWedzichaB. LDodsonA. TAcrylamide is formed in the Maillard reaction. Nature 2002419448
  6. 6. StadlerR. HBlankIVargaNRobertFHauJGuyP. ARobertM. CRiedikerSAcrylamide from Maillard reaction products. Nature 2002419449
  7. 7. YasuharaATanakaYHengelMShibamotoTGas chromatographic investigation of acrylamide formation in browning model systems. Journal of Agricultural and Food Chemistry 2003513999
  8. 8. ZhangYDongYRenYZhangYRapid determination of acrylamide contaminant in conventional fried foods by gas chromatography with electron capture detector. Journal of Chromatography A 20061116209
  9. 9. ZhangYRenYZhaoHZhangYDetermination of acrylamide in Chinese traditional carbohydrate-rich foods using gas chromatography with micro-electron capture detector and isotope dilution liquid chromatography combined with electrospray ionization tandem mass spectrometry. Journal of Analytical Chimica Acta 2007584322
  10. 10. ZamoraRHidalgoF. JContribution of lipid oxidation products to acrylamide formation in model systems. Journal of Agricultural and Food Chemistry 2008566075
  11. 11. CapuanoEOliveiroTAçarO. ÇGökmenVFoglianoVLipid oxidation promotes acrylamide formation in fat-rich model systems. Food Research International 2010431021
  12. 12. KomthongPSuriyaphanOCharoenpanichJDetermination of acrylamide in Thai-conventional snacks from Nong Mon market, Chonburi using GC-MS technique. Food Additives and Contaminants Part B 2012512028
  13. 13. HowardP. HLarge production and priority pollutants. Handbook of Environmental Fate and Exposure Data for Organic Chemicals. Chelsea: Lewis; 1989
  14. 14. TarekeERydbergPKarlssonPErikssonSTörnqvistMAnalysis of acrylamide, a carcinogen formed in heated foodstuffs. Journal of Agricultural Food Chemistry 2002501749985006
  15. 15. CrollB. TArkellG. HHodge RPJ. Residues of acrylamide in water. Water Resource 1974811989993
  16. 16. ShairashiYChromosome aberrations induced by monomeric acrylamide in bone marrow and germ cells of mice. Mutation Research 1978573313324
  17. 17. TilsonH. AThe neurotoxicity of acrylamide. An overview. Neurobehavior Toxicological Teratology 19813455
  18. 18. SicklesD. WBradyS. TTestinoAFriedmanM. AWrennR. WDirect effect of the neurotoxicant acrylamide on kinesin-based microtubule motility. Journal of Neuroscience Research 1996461717
  19. 19. IARCSome Industrial Chemicals. In: IARC Monographs on the Evaluation of Carcinogenic Risk of Chemicals to Humans. France: International Agency for Research on Cancer; 1994
  20. 20. KirkOKirk-Othmer Encyclopedia of Chemical Technology (3rd ed.). New York: John Wiley and Sons; 1978
  21. 21. BachmannMMyersJBezuidenhoutBAcrylamide monomer and peripheral neuropathy in chemical workers. American Journal of Industrial Medicine 1992212217222
  22. 22. BrownLRheadM. MBancroft KCC, Allen N. Case studies of acrylamide pollution resulting from industrial use of acrylamides. Water Pollution Control 1980794507510
  23. 23. SaxN. ILewisR. JHawley’s Condensed Chemical Dictionary (11th ed.). New York: Van Nostrand Reinhold; 1987
  24. 24. Agency for Toxic Substance and Disease Registry (ATSDR)Toxicological profile for acrylamide: draft for public comment. Atlanta: U.S. Public Health Service; 2009
  25. 25. MannsvilleMChemical Products Synopsis, Acrylamide. Mannsville Chemical Products Corporation; 1993
  26. 26. SittigMHandbook of Toxic and Hazardous Chemicals and Carcinogens (2nd ed.). Park Ridge: Noyes; 1985
  27. 27. WongS. STengT. TAhmadaA. LZuhairiANajafpourGTreatment of pulp and paper mill wastewater by polyacrylamide (PAM) in polymer induced flocculation. Journal of Hazardous Materials 2006135378
  28. 28. KirkOKirk-Othmer Encyclopedia of Chemical Technology (3rd ed.) New York: John Wiley and Sons; 1979
  29. 29. MarkH. FGaylordG. NBikalesN. MAcrylamide. Encyclopedia of Polymer Science and Technology. 1962177195
  30. 30. JallageasJ. CArnaudAGalzyPBioconversion of nitriles and their applications. Advances in Biochemical Engineering and Biotechnology 1980141
  31. 31. BouseL. FCarltonJ. BJankP. CUse of polymers for control of spray droplet size. American Society of Agricultural Engineering 1986AA-86-005: 1-18.
  32. 32. DownerR. AWolfT. MChappleA. CHallF. RHazenJ. LCharacterizing the impact of drift management adjuvants on the dose transfer process. Proceeding Fourth International Symposium on Adjuvants Agrochemistry 1995193138
  33. 33. UnderwoodA. KClarkAMackR. EThomasJRobertsJ. RVolgasG. CDry concentrate spray adjuvants. Proceeding Fourth International Symposium on Adjuvants Agrochemistry 1995193391
  34. 34. BrownLRheadM. MBancroft KCC, Allen N. Model studies of the degradation of acrylamide monomer. Water Research 1980147775778
  35. 35. RogachevaS. MIgnatovO. VThe respiratory activity of Rhodococcus rhodochrous M8 cells producing nitrile-hydrolyzing enzymes. Applied Biochemistry and Microbiology 200137282
  36. 36. SmithE. APrunesS. LOehmeF. WEnvironmental degradation of polyacrylamides I. Effects of artificial environmental conditions: temperature, light and pH. Ecotoxicology and Environmental Safety 199635121
  37. 37. SmithE. APrunesS. LOehmeF. WEnvironmental degradation of polyacrylamides II. Effects of environmental exposure. Ecotoxicology and Environmental Safety 19973776
  38. 38. SmithE. AOehmeF. WRapid direct analysis of acrylamide residue in polyacrylamide thickening agents by HPLC. Journal of Chromatographic Science 199331192
  39. 39. Ver Vers LMDetermination of acrylamide monomer in polyacrylamide degradation studies by high-performance liquid chromatography. Journal of Chromatographic Science 199937486
  40. 40. U.S. Environmental Protection Agency (U.S. EPA). Integrated Risk Information System (IRIS) Online. Coversheet for Acrylamide. In: Office of Health and Environmental Assessment, U.S. EPA, Cincinnati; 1994.
  41. 41. ConwayE. JPetersenR. JCollingsworthR. FGracaJ. GCarterJ. WAssessment of the need for and character of limitations on acrylamide and its compounds, Washington DC, US Environmental Protection Agency. (Report prepared for the Offic of Pesticides and Toxic Substances, Contract 68-101979
  42. 42. WeideborgMKällqvistTØdegårdK. ESverdrupL. EVikE. AEnvironmental risk assessment of acrylamide and methyloacrylamide from a grouting agent used in the tunnel construction of Romeriksporten, Norway. Water Research 2001352645
  43. 43. ChangLBruchM. DGriskowitzN. JDentelS. KNMR spectroscopy for determination of cationic polymer concentrations. Water Research 2002362555
  44. 44. LandeS. SBoschS. JHowardP. HDegradation and leaching of acrylamide in soil. Journal of Environmental Quality 19798133
  45. 45. BrownLRheadM. MHillDBancroft KCC. Rapid screening technique utilizing high-performance liquid chromatography for assessing acrylamide contamination in effluents. Analyst 19821071276749754
  46. 46. AsanoYTachibanaMTaniYYamadaHPurification and characterization of amidase which participates in nitrile degradation. Agricultural and Biological Chemistry 1982a46511751181
  47. 47. NagasawaTYamadamHMicrobial transformation of nitriles. Trends Biotechnology 19897153
  48. 48. ShankerRRamakrishnaCSethP. KMicrobial degradation of acrylamide monomer. Archives Microbiology 1990154192
  49. 49. NawazM. SFranklinWCernigliaC. EDegradation of acrylamide by immobilized cells of a Pseudomonas sp. and Xanthomonas maltophilia. Canadian Journal of Microbiology 199339207
  50. 50. NawazM. SKhanA. ASengJ. ELeakeyJ. ESiitonenP. HCernigliaC. EPurification and characterization of an amidase from an acrylamide-degrading Rhodococcus sp. Applied and Environmental Microbiology 1994603343
  51. 51. NawazM. SBilledeauS. MCernigliaC. EInfluence of selected physical parameters on the biodegradation of acrylamide by immobilized cells of Rhodococcus sp. Biodegradation 19989381
  52. 52. WangC. CLeeC. MDenitrification with acrylamide by pure culture of bacteria isolated from acrylonitrile-butadiene-styrene resin manufactured wastewater treatment system. Chemosphere 2001441047
  53. 53. ShukorM. YGusmanizarNAzmiN. AHamidMRamliJShamaanN. ASyedM. AIsolation and characterization of an acrylamide-degrading Bacillus cereus. Journal of Environmental Biology 20093057
  54. 54. ShukorM. YGusmanizarNRamliJShamaanN. AMacCormack WP, Syed MA. Isolation and characterization of an acrylamide-degrading Antarctic bacterium. Journal of Environmental Biology 200930107
  55. 55. ShenS. MWanT. JHwangH. YEnhancement of degradation of acrylamide coupled with salad oil by Pseudomonas aeruginosa DS-4 using incubation periods. Biocatalysis and Agricultural Biotechnology 20121110
  56. 56. AsanoYYasudaTTaniYYamadaHA new enzymatic method of acrylamide production. Agricultural and Biological Chemistry 1982b461183
  57. 57. WamplerD. AEnsignS. APhotoheterotrophic metabolism of acrylamide by a newly isolated strain of Rhodopseudomonas palustris. Applied and Environmental Microbiology 2005715850
  58. 58. WangC. CLeeC. MIsolation of the acrylamide denitrifying bacteria from a wastewater treatment system manufactured with polyacrylonitrile fiber. Current Microbiology 200755339
  59. 59. ChaMChamblissG. HCharacterization of acrylamidase isolated from a newly isolated acrylamide-utilizing bacterium, Rastonia eutropha AUM-01. Current Microbiology 2011622671678
  60. 60. BuranasilpKCharoenpanichJBiodegradation of acrylamide by Enterobacter aerogenes isolated from wastewater in Thailand. Journal of Environmental Sciences 2011233396403
  61. 61. ThanyacharoenUTaniACharoenpanichJIsolation and characterization of Kluyvera georgiana strain with the potential for acrylamide biodegradation. Journal of Environmental Science and Health, Part A 2012471491
  62. 62. ChaMChamblissG. HCloning and sequence analysis of the heat-stable acrylamidase from a newly isolated thermophilic bacterium, Geobacillus thermoglucosidasius AUT-01. Biodegradation 2012DOIs10532-01209557-6.
  63. 63. WakaizumiMYamamotoHFujimotoNOzekiKAcrylamide degradation by filamentous fungi used in food and beverage industries. Journal of Bioscience and Bioengineering 20091085391393
  64. 64. HirrlingerBStolzAKnackmussH. JPurification and properties of an amidase from Rhodococcus erythropolis MP50 which enantioselectively hydrolyzes 2arylpropionamides. Journal of Bacteriology 1996
  65. 65. LiuMJiaoPCaoZ. AAdvances of nitrile hydratase used in bioconversion for acrylamide. Journal of Chemical Industry and Engineering 200152847
  66. 66. GuoX. LDengGXuJWangM. XImmobilization of Rhodococcus sp. AJ270 in alginate capsules and its application in enantioselective biotransformation of trans-2-methyl-3-phenyl-oxiranecarbonitrile and amide. Enzyme and Microbial Technology 200539115
  67. 67. NawazM. SKhanA. ABhattacharayyaDSiitonenP. HCernigliaC. EPhysical, biochemical, and immunological characterization of a thermostable amidase from Klebsiella pneumonia NCTR 1. Journal of Bacteriology 19961782397
  68. 68. Charoenpanich, J, Buranasilp, K, Thanyacharoen, U, Mangkalad, O, Suktanarag, S, Sriwiriyarat, T, & Tani, A. Expression of the recombinant aliphatic amidase from Enterobacter aerogenes in the soluble form and characterization of its activity. In: Summary Book Asian Core Program (2008-2012) on Capacity Building and Development of Microbial Potential and Fermentation Technology Towards New Era (Yamada, M, Leelavatcharamas, V, Xuan, V-T, Bunphanmy, S. eds.) International Exchange Core Program in Yamaguchi University, Japan, ISBN: 978-4-9906826-0-6. November 18-21, Shimonoseki, Yamaguchi, Japan.
  69. 69. LabahnS. KFisherJ. CRobletoE. AYoungM. HMoserD. PMicrobially mediated aerobic and anaerobic degradation of acrylamide in a western United States irrigation canal. Journal of Environmental Quality 201039515631569
  70. 70. CharoenpanichJTaniAProteomic analysis of acrylamide-degradating Enterobacter aerogenes. In: The Science Society of Thailand: Proceedings of the 3rd Biochemistry and Molecular Biology (BMB) international conference From basic to translational researches for a better life, 68April 2011The Empress Convention Centre, Chiangmai, Thailand.
  71. 71. Stams AJMVan Dijk JB, Dijkema C, Plugge CM. Growth of syntrophic propionate-oxidizing bacteria with fumarate in the absence of methanogenic bacteria. Applied and Environmental Microbiology 199359411141119
  72. 72. AnsedeJ. HPellechiaP. JYochD. CMetabolism of acrylate to beta-hydroxypropionate and its role in dimethylsulfoniopropionate lyase induction by a salt marsh sedioment bacterium, Alcaligenes faecalis M3A. Applied and Environmental Microbiology 1999655075
  73. 73. ClarkeP. HThe utilization of amides by microorganisms. In: Payne JW (ed) Microorganisms and nitrogen sources. John Wiley and Sons, New York, 5375621980
  74. 74. PottsJ. RClarkeP. HThe effect of nitrogen limitation on catabolite repression of amidase, histidase and urocanase in Pseudomonas aeruginosa. Journal of General Microbiology 197693377
  75. 75. SmythP. FClarkeP. HCatabolite repression of Pseudomonas aeruginosa amidase: The effect of carbon source on amidase synthesis. Journal of General Microbiology 19759081
  76. 76. JohnstonJ. JBordenR. CBarlazM. AAnaerobic biodegradation of alkylbenzenes and trichloroethylene in aquifer sediment down gradient of a sanitary landfill. Journal of Contaminants Hydrolysis 199623263
  77. 77. MalloryL. MYukC. SAlexanderMAlternative prey: a mechanism for elimination of bacterial species by protozoa. Applied and Environmental Microbiology 1983461073
  78. 78. MurakamiYAlexanderMDestruction and formation of toxins by one bacterial species affect biodegradation by a second species. Biotechnology and Bioenergetics 198933832
  79. 79. GaoMWangD. XZhengQ. YHuangZ. TWangM. XRemarkable electronic and steric effects in the nitrile biotransformations for the preparation of enantiopure functionalized carboxylic acids and amides: implication for an unsaturated carbon-carbon bond binding domain of the amidase. Journal of Organic Chemistry 2007726060
  80. 80. SongLWangMShiJXueZWangM. XQianSHigh resolution X-ray molecular structure of the nitrile hydratase from Rhodococcus erythropolis AJ270 reveals posttranslational oxidation of two cysteines into sulfinic acids and a novel biocatalytic nitrile hydration mechanism. Biochemical and Biophysical Research Communication 2007362319
  81. 81. WangJ. YWangD. XPanJHuangZ. TWangM. XNitrile and amide biotransformations for the synthesis of enantiomerically pure 3arylaziridine-2-carboxamide derivatives and their stereospecific ring-opening reactions. Journal of Organic Chemistry 2007
  82. 82. EndoINojiriMTsujimuraMNakasakoMNagashimaSYohdaMOdakaMFe-type nitrile hydratase. Journal of Inorganic Biochemistry 200183247
  83. 83. YamadaHKobayashiMNitrile hydratase and its application to industrial production of acrylamide. Bioscience Biotechnology and Biochemistry 1996601391
  84. 84. MayauxJ. FCerbelaudESoubrierFYehPBlancheFPetreDPurification, cloning, and primary structure of a new enantionmer-selective amidase from a Rhodococcus strain: structural evidence for a conserved genetic coupling with nitrile hydratase. Journal of Bacteriology 19911736694
  85. 85. WangC. CLeeC. MWuA. SAcrylic acid removal from synthetic wastewater and industrial wastewater using Ralstonia solanacearum and Acidovorax avenae isolated from a wastewater treatment system manufactured with polyacrylonitrile fiber. Water Science and Technology 2009601130113016
  86. 86. StraathofA. JSieSFrancoT. TWielenL. AFeasibility of acrylic acid production by fermentation. Applied Microbiology and Biotechnology 200567727
  87. 87. SchomburgDSalzmannMEnzyme Handbook 4. Berlin, Springer-Verlag; 1991
  88. 88. BorkPKooninE. VA new family of carbon-nitrogen hydrolases. Protein Science 199431344
  89. 89. NovoCFarnaudSTataRClementeABrownP. RSupport for a three dimensional structure predicting a Cys-Glu-Lys catalytic triad for Pseudomonas aeruginosa amidase comes from site-directed mutagenesis and mutations altering substrate specificity. Biochemical Journal 2002365731
  90. 90. AndradeJKarmaliACarrondoM. AFrazaoCStructure of amidase from Pseudomonas aeruginosa showing a trapped acyl transfer reaction intermediate state. Journal of Biology and Chemical 200728219598
  91. 91. SuktanaragSCharoenpanichJSequence analysis and structural modeling of an amidase from a novel acrylamide-degrading bacterium Enterobacter aerogenes. The 13th FAOBMB Congress, November 25-29, 2012Bangkok, Thailand.
  92. 92. ChinK. HTsaiY. DChanN. LHuangK. FWang AHJ, Chou SH. The crystal structure of XC1258 from Xanthomonas campestris: A putative prokaryotic Nit protein with an arsenic adduct in the active site. Proteins 200769665
  93. 93. KimaniS. WAgarkarV. BCowanD. ASayed MFR, Sewell BT. Structure of an aliphatic amidase from Geobacillus pallidus RAPc8. Acta Crystallography 2007631048
  94. 94. KobayashiMFujiwaraYGodaMKomedaHShimizuSIdentification of active sites in amidase: Evolutionary relationship between amide bond-and peptide bond-cleaving enzymes. Proceedings of the National Academy of Science USA 19979411986991
  95. 95. NovoCTataRClementeABrownP. RPseudomonas aeruginosa aliphatic amidase is related to the nitrilase/cyanide hydratase enzyme family and Cys166 is predicted to be the active site nucleophile of the catalytic mechanism. FEBS Letters 1995367275

Written By

Jittima Charoenpanich

Submitted: July 4th, 2012 Published: October 2nd, 2013