Open access peer-reviewed chapter

Missense Mutation in AR-CGD

Written By

M. Yavuz Köker and Hüseyin Avcilar

Submitted: May 15th, 2011 Reviewed: November 11th, 2011 Published: October 12th, 2012

DOI: 10.5772/35758

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1. Introduction

Chronic granulomatous disease (CGD) is an inherited disorder of the innate immune system characterized by impairment of intracellular microbicidal activity of phagocytes. Mutations in one of four known nicotinamide adenine dinucleotide phosphate (NADPH) -oxidase components preclude generation of superoxide and related antimicrobial oxidants, leading to the phenotype of CGD. Defects in gp91-phox, encoded by CYBBgene, lead to X-linked CGD and have been reported to be responsible for approximately 65% of all CGD cases. The autosomal gene in CGD are CYBA, encoding p22-phox, NCF2, encoding p67-phox, NCF1, encoding p47-phox, and NCF4, encoding p40-phox (figure 1) (1,2). The mutation in thesegenes, respectively, abolishes the activity of the oxidase and leads to autosomal recessive chronic granulomatous disease (AR-CGD) which is approximately 35% of all CGD cases (table 1).

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2. Phenotype- genotype correlation in CGD

Identification of specific mutations in CGD patients may help to clarify some of the variability in clinical severity seen in this disorder and shows genotype-phenotype correlation. In general, X-CGD patients follow a more severe clinical course than patients with an AR-CGD and exhibit in the first years of life. AR-CGD patients follow a milder clinical course, especially p47-phox defect, and mostly seen in first and second decade of life. AR-CGD patients with missense mutations usually exhibit a mild clinical course, associated with a residual activity of p47-phox and also p22 and p67-phoxs. However, the level of superoxide generation does not always correlate with the clinical course. Some patients suffer from severe and recurrent infections despite having neutrophils with 10–20% of normal oxidase activity (1). Within our study with 40 AR-CGD families, we could not define a direct correlation between the molecular defect and the clinical course of the disease. Either truncations (nonsense and frameshift mutations) or missense mutations could have resulted in severe influence on phenotype.

Figure 1.

NADPH oxidase enzyme subunits and complex in activation phase.

Table 1.

The molecular characteristic of NADPH Oxidase components.

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3. CYBA(cytochrome balfa chain)gene

Cytochrome bis comprised of a light a-chain and a heavy b-chain. This gene encodes the light, alpha subunit which has been proposed as a primary component of the microbicidal oxidase system of phagocytes. Mutations in this gene are associated with AR-CGD that is characterized by the failure of activated phagocytes to generate superoxide, which is important for the microbicidal activity of these cells. http://www.genecards.org/cgi-bin/carddisp.plgene=CYBA.

In about 5% of the CGD patients, the disease is caused by mutations in the cytochrome balfa chain (CYBA)gene. The CYBAgene encoding p22-phox which contains 195 amino acid, is localized on chromosome 16q24, has a size of about 8,5 and contains six exons and trans-membrane and proline rich domains [3, 4]. CYBAgene encoding p22-phox has 19 different missense mutations in 65 mutated alleles and has more missense mutation than other NADPH oxidase subunit genes (table 2a) (figure 2). Mutations in the CYBAgene have been updated by [Roos et al., (2)] and are reviewed in Human Gene Mutation Database, (HGMD; http://www.hgmd.cf.ac.uk/ac/all.php).

P22-phox has a key role in the interaction of NADPH-oxidase subunits and any difference in amino acid pattern of this phox protein may change the globular conformation of this protein due to the difference in the electrophoretic characteristic of new amino acid, which prevents the complex formation with other subunits.

Figure 2.

Missense mutations inCYBAgene, encoding p22-phox;CYBAgene contains 6 exons. P22-phox contains trans-membrane and proline rich domains. Missense mutation points inCYBAgene and change in encoded 195 aa. of p22-phox represented in thefigure (2).

Missense mutation points may have an important role in the interaction with other subunits, so the amino acid change in that regions may change the property of interactions and prevents or decreases the complex formation, so the activity of NADPH oxidase was abolished.

Total numbers of alleles which have missense mutations are 65 of 173 mutated alleles in CYBAgene and the percentage of missense mutations in that mutated alleles are %37,5 (table 2a) (2).Percentage of missense and all mutations of CYBAgene in the overall mutations of AR-CGD is %6.3 and %16.8, respectively (table 3 and 4). Most prevalent missense mutations points in CYBAgene are c.70G>A, c.268C>T and c.354C>A which cause p.Gly24Arg, p.Arg90Trp and p.Ser118Arg in p22-phox, respectively (table 5).

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4. NCF1(Neutrophil Cytosolic Factor 1) gene

In about 25% of the CGD patients, the disease is caused by mutations in the neutrophil cytosolic factor 1 (NCF1) gene on chromosome 7q11.23, which encodes p47phox, one of the structural components of the NADPH oxidase and has a size of about 40 kb and contains 11 exons (5, 6). The protein encoded by this gene is a 47 kDa cytosolic subunit of neutrophil NADPH oxidase and is required for activation of the latent NADPH oxidase and contains 390 amino acids and PX, SH3a, SH3b and polybasic domains (figure 3).

Figure 3.

Missense mutations inNCF1gene, encoding p47-phox;NCF1gene contains 11 exons. p47-phox contains PX, SH3a, SH3b and polybasic domains. Mutation points inNCF1gene and change in encoded 390 aa. of p47-phox represented in thefigure (2).

A very common mutation found in these patients is a GT deletion in a GTGTrepeat sequence at the beginning of exon 2 of NCF1 (c.75_76delGT) gene (5, 7). NCF1gene encoding p47-phox has only 4 different missense mutation in 6 alleles (table 2b) (figure 3) (2). P47-phox has an important role in the interaction of cytoplasmic NADPH-oxidase subunits and any difference in amino acid pattern of this phox protein may abolish the complex formation with other subunits.

Total numbers of alleles which have missense mutations are 6 of 63 (other than delta-GT mutation in exon 2, in more than 620 alleles) mutated alleles in NCF1gene and the percentage of missense mutations in that mutated alleles are %9,5. The percentage of missense and all mutations (including delta-GT mutation) of NCF1gene in the overall mutations of AR-CGD is %0.6 and %66.4, respectively (table 3 and 4). So, this high percentage due to the high number of delta-GT mutation in exon 2 of NCF1gene and is more than all the mutations in AR-CGD (table 5). This deletion points is hot-spot mutation region for NCF1gene.

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5. NCF2(Neutrophil Cytosolic Factor 2) gene

The neutrophil cytosolic factor 2 (NCF2)gene encoding p67-phox is localized on chromosome 1q25, has a size of about 40 kb and contains 16 exons and TRP1-4, AD, SH3a, PB1 and SH3b domains (figure 4) (7, 8). NCF2gene, encoding p67-phox, has 41 different missense mutations in 171 mutated alleles (table 2c) (2, 4, 5, 6). Mutations in the NCF2gene have been published by [Roos et al., (2)] and are reviewed in Human Gene Mutation Database, (HGMD; http://www.hgmd.cf.ac.uk/ac/all.php). P67-phox has a major role in the interaction of NADPH-oxidase subunits in cytoplasm and any difference in amino acid pattern of this phox protein may prevent the complex formation with other subunits (figure 1).

Figure 4.

Missense mutations in NCF2 gene, encoding p67-phox;NCF2gene contains 16 exons. P67-phox contains TRP1-4, AD, SH3a, PB1 and SH3b domains. Mutation points inNCF2gene and change in encoded 526 aa. of p67-phox represented in thefigure (2).

Total numbers of mutated alleles leading AR-CGD in NCF2gene are 171 and 41 of them are missense and percentage of missense mutations in that mutated alleles are %24 (table 2c) (2).Percentage of missense and all mutations of NCF2gene in the overall mutations of AR-CGD is %4 and %16.6, respectively (table 3 and 4). Most prevalent missense mutations points in NCF2gene is c.279C>G which causes p.Asp93Glu in p67-phox (table 5).

Table 2.

(a) Missense Mutation in CYBAgene. (b) Missense Mutation in NCF1gene. (c) Missense Mutation in NCF2gene.

Table 3.

Distribution of number and percentage of missense and all mutations in genes (CYBA, NCF1, NCF2and NCF4) of AR-CGD.

Table 4.

Number and percentage of different missense mutations in genes (CYBA, NCF1, NCF2and NCF4) of AR-CGD.

Table 5.

Most prevalent missense mutation in the genes of AR-CGD.

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6. NCF4(Neutrophil Cytosolic Factor 2) gene

NCF4gene encoding p40-phox with 339 amino acids is localized on chromosome 22q13.1 has a size of about 4,4 kb and contains 10 exons. P40-phox interacts primarily with p67-phox. Up to know, the first mutation in NCF4gene was founded in a family with compound heterozygote mutations and one of the mutations was a missense with c.314G>A in one allele, which causes change in p.Arg105Gln amino acid in the structure of p40-phox (2, 9).

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7. Conclusion

19 different alleles in CYBAgene, 4 different alleles in NCF1gene, 17 different alleles in NCF2 gene and one allele in NCF4gene have missense mutations which cause change in amino acid patterns of NADPH oxidase subunits and results in AR-CGD. The percentage of missense mutations in the overall mutations of AR-CGD is %11 (table 3). One of the most prevalent missense mutations in AR-CGD is in CYBAgenewith c.70G>A, in 14 alleles of 9 families, which causes p.Gly24Arg in p22-phox (table 5).

In p22-phox the first interaction with p67-phox occur in B part (domain) which is located between 81-91 amino acids in p22-phox. There are 4 different missense mutations (in 21 alleles of CYBAgene) change amino acid (arginine) at position 90. So, this position is highly susceptible to any conformational changes which may prevent the interaction with p67-phox. So, the change in the molecular structure of this part may abolish the stability and function of p22-phox and latent NADPH oxidase could not be activated leading to AR-CGD. P22-phox has more different missense mutation than other NADPH oxidase components. The ratio of the number of different missense mutation and the number of amino acid in the chain is approximately 19/195 (%9.74). The different missense mutation to overall amino acid chain length in p67-phox is 17/526 (%3.23). But, the ratio in p47-phox is 4/390 (%1). This result shows that p67-phox has 3 times and p22-phox has approximately 10 times high incidence of different missense mutations than p47-phox in their primary amino acid structure. The underlying reason for this may be the highly specific interaction and function of p22-phox which is vulnerable to any change in the globular structure of protein.

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Acknowledgement

This study is kindly supported by TÜBİTAK with project number 110S252.

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Written By

M. Yavuz Köker and Hüseyin Avcilar

Submitted: May 15th, 2011 Reviewed: November 11th, 2011 Published: October 12th, 2012