Vitamins are important micronutrients that are often precursors to enzymes, which all living cells require to perform biochemical reactions. However, humans cannot produce many vitamins, so they have to be externally obtained. Using vitamin‐producing microorganisms could be an organic and marketable solution to using pseudo‐vitamins that are chemically produced, and could allow for the production of foods with higher levels of vitamins that could reduce unwanted side effects. Probiotic bacteria, as well as commensal bacteria found in the human gut, such as Lactobacillus and Bifidobacterium, can de novo synthesize and supply vitamins to human body. In humans, members of the gut microbiota are able to synthesize vitamin K, as well as most of the water‐soluble B vitamins, such as cobalamin, folates, pyridoxine, riboflavin, and thiamine.
Vitamins are typically categorized as fat‐soluble vitamins, which includes vitamins A, D, E, and K, or as water‐soluble vitamins, which includes vitamin C, biotin (vitamin H or B7), and a series of B vitamins—thiamin (B1), riboflavin (B2), niacin (B3), pantothenic acid (B5), pyridoxine (B6), folic acid (B11), and cobalamin (B12). While fat‐soluble vitamins act as important elements of cell membranes, water‐soluble vitamins serve as coenzymes that typically transport specific chemical groups . Humans are incapable of synthesizing most vitamins and they consequently have to be obtained exogenously. The use of vitamin‐producing microorganisms might represent a more natural and consumer‐friendly alternative to fortification using chemically synthesized pseudo‐vitamins.
The biochemical pathways involved in B‐vitamin biosynthesis by food microorganisms were previously described in detail . Many prokaryotes need water‐soluble vitamins for nutritional purposes , but also typically need them for biosynthetic processes. The ability of particular microorganisms to produce B vitamins could supplant the expensive chemical production of these vitamins to enrich food or be improved for in situ fortification of fermented foods. Much research has been conducted in recent years to elucidate the biosynthetic pathways of these vitamins in a number of microorganisms.
Probiotic bacteria positively impact the immune system and the composition and functioning of the gut microbiota . Furthermore, the production of vitamins has resulted in many healthy benefits to the host. Probiotic bacteria, mostly belonging to the genera
The production of B‐vitamins, especially folate and riboflavin (B2), by probiotic bacteria has been extensively researched as described in a recent review [7, 8]. Several lactic acid bacteria (LAB) species (e.g.,
This review focused on riboflavin, folic acid, and cobalamin, three of the water‐soluble B vitamins whose biosynthetic pathways were inextricably linked, briefly covering their physiological functions and dietary sources before concentrating on novel overproduction strategies in probiotics.
2. Riboflavin biosynthesis
In contrast to many plants, fungi, and bacteria, humans cannot produce riboflavin or vitamin B2, and thus require it as a dietary supplement. Riboflavin is available as a dietary source and is also produced by the microflora of the large intestine [6, 14]. Riboflavin (vitamin B2) plays an essential role in cellular metabolism, as it is the precursor of the coenzymes flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), which both act as hydrogen carriers in many biological redox reactions.
Riboflavin is synthesized by many bacteria and its biosynthetic pathway has been studied extensively in
The ability of some bacteria and fungi to overproduce riboflavin has been harnessed for industrial production. Such commercial producers include the ascomycetes
It has been reported that fermentation of cow milk with
Species and/or strain‐specific traits in LAB provided genetic information for riboflavin biosynthesis. Several of the sequenced members of LAB possessed similar abilities to produce riboflavin, as suggested by comparative genome analysis, but an interrupted
3. Folate biosynthesis by human gut commensals
Folic acid, also known as vitamin B11, is a dietary necessity for humans, because it is used in several metabolic reactions, such as the biosynthesis of the building blocks of DNA and RNA, the nucleotides. It is recommended that adults take 200 μg daily, but pregnant women are encouraged to take a double dose daily, as folic acid could thwart neural‐tube defects in newborns . Low folic acid has been linked to high homocysteine levels in the blood, which could lead to coronary diseases [31, 32]. It has also been shown to protect against some forms of cancer . Folate is conspicuously absent in many food products and is considered an essential additive to the general diet.
Folates are comprised of a mono‐ or polyglutamyl conjugate and these compounds were named after the number of glutamyl residues (PteGlu
Folate biosynthesis continues with the formation of a C–N bond joining DHPPP to pABA. This condensation reaction, catalyzed by dihydropteroate synthase, yields 7,8‐dihydropteroate (DHP). DHP is glutamylated by dihydrofolate synthase, resulting in dihydrofolate (DHF). It is then reduced by DHF reductase to the biologically active cofactor tetrahydrofolate (THF) and subjected to the addition of multiple glutamate moieties by folylpolyglutamate synthase to yield THF‐polyglutamate. Polyglutamilation may also take place before the occurrence of the reduction step, being catalyzed by DHF synthase or, in many bacteria, by a bifunctional enzyme that is responsible for both DHF synthase and folylpolyglutamate synthase activities .
However, although all available complete bifidobacterial genomes are expected to specify aminodeoxychorismate synthase, a gene specifying a putative 4‐amino‐4‐deoxychorismate lyase can only be found on the genome of
Lactobacilli are also typical human gut commensals and were recently investigated to discover if they could serve as possible folate producers . Lactobacilli from various fermented foods have been investigated as starter cultures for the manufacturing of folate‐fortified dairy products, while lactobacilli isolated from the human gut have been explored as folate‐producing probiotics [39–42]. The availability of genome sequences of various lactobacilli provided an important contribution to the genetics underlying folate biosynthesis in this group of microorganisms . For example, lactobacilli did not appear to harbor the genetic determinants for de novo pABA synthesis, with the exception of
Currently, the strains of
4. Vitamin B12 biosynthesis
Vitamin B12, otherwise known as cobalamin, is the biggest and most intricate vitamin. Cobalamin describes a cluster of cobalt‐containing compounds (corrinoids) that have a lower axial ligand, which holds the cobalt‐coordinated nucleotide (5, 6‐dimethylbenzimidazole) as a base. Although humans only use vitamin B12 for two enzymatic activities, it is still an important dietary supplement. (R)‐methyl‐malonyl‐CoA mutase assists in the metabolism of propionyl‐CoA, which compounds such as valine, thymine, methionine, and odd‐chain fatty acids produce when broken down. This ado‐cobalamin‐dependent enzyme catalyzes the rearrangement of propionyl‐CoA following its carboxylation and epimerization to succinyl‐CoA, which then goes through the citric acid cycle. Methionine synthase needs vitamin B12 in the form of methylcobalamin. Using 5‐methyltetrahydrofolate as a methyl donor, this enzyme methylates homocysteine to form methionine .
Humans cannot synthesize vitamin B12, and, thus must obtain it from organisms that can. Only a limited number of bacteria are known to produce vitamin B12, three of which—
Cobalamin has the most complex structure of all the vitamins synthesized by bacteria requiring about 30 genes for its biosynthesis. Most of the work in characterizing cobalamin biosynthesis has been performed in
Genes encoding enzymes contributing to the oxygen‐dependent pathway have been given the prefix
LAB are traditionally known as auxotrophic for cobalamin and are generally used for the biological analysis of this vitamin. Recently, however, cobalamins were identified in
The complete genome of
Other strains of genus
5. Biosynthesis of other B‐group vitamins
Thiamine (vitamin B1) is a coenzyme in the pentose phosphate pathway that is required to synthesize fatty acids, steroids, nucleic acids, and the aromatic amino acid precursors into various neurotransmitters and other bioactive compounds essential for brain function . Beyond its role as a necessary cofactor in the folate cycle, vitamin B6 (pyridoxine) also plays an important role in amino acid metabolism, which makes it a rate‐limiting cofactor in the synthesis of neurotransmitters such as dopamine, serotonin, gamma‐aminobutyric acid (GABA), noradrenaline, and the hormone melatonin .
LAB fermentation in yogurt, cheese, and other fermented products was shown to result in increased levels of riboflavin, folate, vitamin B12, niacin, and pyridoxine [65, 66]. Soy fermentation with
6. Biosynthesis of vitamin K
Vitamin K serves as a cofactor for the enzyme that converts specific glutamyl residues in a few proteins to g‐carboxyglutamyl (Gla) residues, aiding in the process. Humans obtain the daily nutritional requirement for vitamin K through the dietary phylloquinone that exists in plants, and, to some extent, through bacterially produced polyisoprenyl‐containing compounds called menaquinones created in the human gut . LAB were examined for their ability to produce quinone compounds, as vitamin K occurred naturally in two forms, namely, K1 (phylloquinone) in green plants, and K2 (menaquinones) in animals and some bacteria .
The use of vitamin‐producing strains provided a new perspective on the specific uses of probiotics. Many food‐grade bacteria overproduce B vitamins, including riboflavin (vitamin B2), folate (vitamin B11), and cyanocobalamine (vitamin B12), which could allow them to organically enrich raw food materials like soy, milk, meat, and vegetables with B vitamins, preventing the need for additives. Thus, the food industry could take advantage of these novel and efficient vitamin‐producing strains to add nutritional value to fermented products and save money in the process. Notably, vitamin metabolism pathways were shown in genes that specified the biosynthetic enzymes for riboflavin, cobalamin, and folate production. It is increasingly possible to identify potential vitamin‐producing strains and interpret the intertwined mechanisms for their biosynthesis, because of the expanding availability of genome sequences, which could be used to expand the vitamin‐producing capacities of the human gut.
This project was funded by the International Science & Technology Cooperation Program of China (2013DFA32330), the National Natural Science Foundation of China (No. 31071513, No. 31271821), the Natural Science Foundation of Zhejiang Province (No. LY16C200002), the National High Technology Research and Development Program (“863” Program) of China (2012AA022105B), the National Research Foundation for the Doctoral Program of Higher Education (20133326110005), and the Science Foundation of the Zhejiang Education Department (No. Y201534497).