Isolation and Characterization of Protease-Deficient Mutants of Vibrio cholerae

Schneider, D.R.; Parker, C D

doi:10.1093/infdis/138.2.143

Cited by 49 publications

(26 citation statements)

References 38 publications

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“…Protease-deficient mutants of V. cholerae are reported to be less virulent (35). The most well-studied protease in cholera is hemagglutinin protease.…”

Section: Discussionmentioning

confidence: 99%

Enterotoxigenicity of Mature 45-Kilodalton and Processed 35-Kilodalton Forms of Hemagglutinin Protease Purified from a Cholera Toxin Gene-Negative Vibrio cholerae Non-O1, Non-O139 Strain

Ghosh¹,

Saha²,

Hoque³

et al. 2006

Infect Immun

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Cholera toxin gene-negativeVibrio cholerae non-O1, non-O139 strains are an intriguing group that can cause typical cholera-like disease but do not produce cholera toxin (CT) (23). The virulence factor in V. cholerae non-O1, non-O139 strains that do not produce cholera toxin and yet cause symptoms similar to cholera has not yet been identified (6). It was found that none of the previously described virulence-related genes like the RTX (repeats in toxin) toxin gene cluster, the hlyA gene coding for hemolysin, the mshA gene for mannose-sensitive hemagglutinin pilus, and the stn gene encoding a heat-stable enterotoxin of non-O1 vibrios (NAG-ST) was specifically associated with the pathogenesis of TCP Ϫ CTX Ϫ non-O1, non-O139 strains that colonized mice and caused fluid accumulation in rabbits (6). In the above-mentioned study, the role of the hapA gene (coding for hemagglutinin protease [HAP]) in the disease process was not investigated. Studies by Benítez et al. with vaccine strain 638, which bears a core deletion of the ctx gene and an insertional mutation of the hapA gene, showed fewer symptoms and a decrease in diarrhea in volunteers compared to those in previous studies using only the core deletion strain (2). In addition, strain 638 induced lower levels of interleukin-8 (IL-8) than its parent wild-type strain did in HT29 cells (2, 30). Cholera vaccine strain CVD 110, a mutant of El Tor Ogawa strain E7946, which is deleted of all known virulence genes except hap, gives rise to a highly inflammatory reaction (36). Concentrated proteins derived from the culture supernatant of CVD 110 caused morphological changes in cultured MDCK-1 epithelial cells and altered their arrangement of filamentous actin and zonula occludens-associated protein (ZO-1) (38). The drastic morphological changes were inhibited by Zincov, a specific bacterial metalloprotease inhibitor. After fast-performance liquid chromatography, the cytotoxic fractions of the sample showed two visible bands with molecular masses of approximately 34 and 32 kDa. Microsequencing of these proteins revealed that they were in fact HAP. Culture supernatants prepared from the reactogenic strains of V. cholerae cause a decrease in the transcellular epithelial resistance of T84 intestinal cells (21). This decrease correlates with the presence of HAP but not with the presence of other potential accessory toxins or proteases (21). Microarray studies of the global transcription pattern of V. cholerae grown in vivo in the rabbit ileal loop (RIL) model have shown that the hemagglutinin protease (hap) gene is one of the 12 genes that belong to the pathogenesis functional group and are highly expressed compared to their levels of expression under laboratory conditions (29). V. cholerae O1 classical and El Tor biotypes as well as non-O1 serotypes produce HAP (3). The secreted HAP has been purified, cloned, and sequenced (9, 14). The hap gene consists of 1,827 nucleotides with a predicted molecular mass of 69.3 kDa. This is larger than the molecular mass of the purified HAP (3...

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“…Protease-deficient mutants of V. cholerae are reported to be less virulent (35). The most well-studied protease in cholera is hemagglutinin protease.…”

Section: Discussionmentioning

confidence: 99%

Enterotoxigenicity of Mature 45-Kilodalton and Processed 35-Kilodalton Forms of Hemagglutinin Protease Purified from a Cholera Toxin Gene-Negative Vibrio cholerae Non-O1, Non-O139 Strain

Ghosh¹,

Saha²,

Hoque³

et al. 2006

Infect Immun

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“…Protease-deficient mutants of V. cholerae are k n o w n to be less virulent (16). Protease-deficient mutants of V. cholerae are k n o w n to be less virulent (16).…”

Section: Introductionmentioning

confidence: 99%

The protease from Vibrio Cholerae nicks arginine at position 192 from the N-terminus of the heat-labile enterotoxin a subunit from enterotoxigenic Escherichia Coli

et al. 1992

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It was examined where a protease purified from Vibrio cholerae might nick the heat-labile enterotoxin (LT) A subunit from enterotoxigenic Escherichia coli. LT was digested by the protease and contained a fragment which had the same mobility on SDS-PAGE as that of the A1 fragment of LT digested by trypsin. The biological activity of LT by this protease was also identical to that of LT by trypsin. The amino acid sequence of the N-terminus of the A2-like fragment was Thr-Ser-Thr-Gly, which corresponded to the sequence from 193 to 196 of the A subunit. These data suggest that this protease, like trypsin, nicks arginine at position 192 from the N-terminus of the A subunit and that the biological activation of LT by this protease is similar to that by trypsin.

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“…After Sephadex G100 partial purification, this yield increased to 436IU/L. As found by other groups interested in the exoenzymes of V. choZerae (Schick and Zilg, 1977;Parker, 1978 and1982), it was apparent that a high degree of purification had to be sacrificed in order to maintain high yields of mucinase.…”

Section: Discussionmentioning

confidence: 77%

Studies on the Vibrio cholerae mucinase complex. I. Enzymic activities associated with the complex325

Stewart‐Tull

Ollar

Scobie

1986

Journal of Medical Microbiology

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Isolation and Characterization of Protease-Deficient Mutants of Vibrio cholerae

Cited by 49 publications

References 38 publications

Enterotoxigenicity of Mature 45-Kilodalton and Processed 35-Kilodalton Forms of Hemagglutinin Protease Purified from a Cholera Toxin Gene-Negative Vibrio cholerae Non-O1, Non-O139 Strain

Enterotoxigenicity of Mature 45-Kilodalton and Processed 35-Kilodalton Forms of Hemagglutinin Protease Purified from a Cholera Toxin Gene-Negative Vibrio cholerae Non-O1, Non-O139 Strain

The protease from Vibrio Cholerae nicks arginine at position 192 from the N-terminus of the heat-labile enterotoxin a subunit from enterotoxigenic Escherichia Coli

Studies on the Vibrio cholerae mucinase complex. I. Enzymic activities associated with the complex325

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