Cloning and characterization of a gene encoding a new thermostable hemolysin from Vibrio parahaemolyticus

Taniguchi, Hatsumi; Kubomura, Shigeo; Hirano, Hideaki; Mizue, Kumiko; Ogawa, Midori; Mizuguchi, Yasuo

doi:10.1016/0378-1097(90)90020-q

Cited by 17 publications

(18 citation statements)

References 13 publications

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“…2002). However, this is in agreement with the V. parahaemolyticus δ ‐VPH, which does not contain any features typical of a signal sequence at its amino‐terminus (Taniguchi et al. 1990).…”

Section: Introductionsupporting

confidence: 79%

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Haemolysins in Vibrio species

2005

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Many Vibrio species are pathogenic to humans, and/or marine vertebrates and invertebrates. The pathogenic species produce various virulence factors including enterotoxin, haemolysin, cytotoxin, protease, lipase, phospholipase, siderophore, adhesive factor and/or haemagglutinins. Haemolysin, which is an exotoxin that lyses erythrocyte membranes with the liberation of haemoglobin, is arguably the most widely distributed toxin among pathogenic vibrios and exerts various roles in the infection process. Haemolysins act on erythrocytes membranes thus lysing the cells which leads to the freeing up of the iron-binding proteins namely haemoglobin, transferrin and lactoferrin. This iron can then be picked up by various siderophores, and is subsequently taken up through receptors in the cell membrane. In many cases, the pore-forming activity of haemolysin is not restricted to erythrocytes, but extends to a wide range of other cell types including mast cells, neutrophils, and polymorphonuclear cells, and enhances virulence by causing tissue damage. There are four representative haemolysin families in Vibrio spp., including the TDH (thermostable direct haemolysin) family, the HlyA (E1 Tor haemolysin) family, the TLH (thermolabile haemolysin) family and the d-VPH (thermostable haemolysin) family. Some haemolysins, for example, TDH of Vibrio parahaemolyticus and HlyA of Vibrio cholerae have been studied extensively, and are closely associated with virulence. However, the role of some haemolysins, e.g. TLH and d-VPH are unclear, and await the outcome of further research.

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Section: Introductionsupporting

confidence: 79%

“…The gene encodes a polypeptide of 203 amino acid residues. Interestingly, the nucleotide and predicted amino acid sequence differ from those of the tdh , trh and tlh genes, or any other haemolysins of V. parahaemolyticus (Taniguchi et al. 1990).…”

Section: Introductionmentioning

confidence: 99%

Haemolysins in Vibrio species

2005

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“…However, identifying the pathogenic serovars of V. parahaemolyticus by use of only these two toxins is not sufficiently accurate, as other hemolysins may take part in the pathogenicity of V. parahaemolyticus. Thermolabile hemolysin (TLH), a toxin encoded by the tlh gene of V. parahaemolyticus and present in almost every clinical and environmental V. parahaemolyticus strain (14, 37), has been suggested as a promising target for pathogen detection (30,35,44). Although TLH has hemolytic activity and can lyse red blood cells, its cytotoxic and biochemical mechanisms of action are still not clearly understood (3,26,31).…”

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confidence: 99%

Screening for a Single-Chain Variable-Fragment Antibody That Can Effectively Neutralize the Cytotoxicity of the Vibrio parahaemolyticus Thermolabile Hemolysin

Wang

Sui

et al. 2012

Appl Environ Microbiol

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bVibrio parahaemolyticus is a halophilic bacterium that is widely distributed in water resources. The bacterium causes lethal food-borne diseases and poses a serious threat to human and animal health all over the world. The major pathogenic factor of V. parahaemolyticus is thermolabile hemolysin (TLH), encoded by the tlh gene, but its toxicity mechanisms are unknown. A highaffinity antibody that can neutralize TLH activity effectively is not available. In this study, we successfully expressed and purified the TLH antigen and discovered a high-affinity antibody to TLH, named scFv-LA3, by phage display screening. Cytotoxicity analysis showed that scFv-LA3 has strong neutralization effects on TLH-induced cell toxicity. Vibrio parahaemolyticus, a Gram-negative motile bacterium that inhabits marine and estuarine environments throughout the world (7,23,36), is a major food-borne pathogen that causes life-threatening diseases in humans through the consumption of raw or undercooked seafood (5,21,34 It is well known that V. parahaemolyticus contains many different kinds of toxins, such as thermostable direct hemolysin (TDH), TDH-related hemolysin (TRH), and some noncharacterized proteins. TDH is considered one of the major virulence factors of V. parahaemolyticus, and its function has been well characterized and discussed (25). TRH is another hemolysin of V. parahaemolyticus that can also lyse red blood cells, and it has high sequence homology with TDH (24, 42). However, identifying the pathogenic serovars of V. parahaemolyticus by use of only these two toxins is not sufficiently accurate, as other hemolysins may take part in the pathogenicity of V. parahaemolyticus. Thermolabile hemolysin (TLH), a toxin encoded by the tlh gene of V. parahaemolyticus and present in almost every clinical and environmental V. parahaemolyticus strain (14, 37), has been suggested as a promising target for pathogen detection (30,35,44). Although TLH has hemolytic activity and can lyse red blood cells, its cytotoxic and biochemical mechanisms of action are still not clearly understood (3,26,31). Since TLH may be as important as TDH and TRH (6), it is necessary to investigate its function during the process of infection.Single-chain variable-fragment (scFv) antibody generation is a versatile technology for generating antibodies that are specific for a given antigen (40). It has also been used for selective molecular targeting in cancer research for conditions such as lymphatic invasion vessels, colon cancer, and hepatocarcinoma (27,29,33,43). Furthermore, scFv antibody generation has been used extensively to generate ligands for detecting pathogenic germs in vitro and in vivo (8,22,38,39). Compared to polyclonal antibodies or hybridoma technology, scFv antibodies can easily be manipulated genetically to improve their specificity and affinity, reducing production costs. In addition, they can be fused with molecular markers for immunological detection of pathogenic bacteria (10, 28). scFv antibody generation has also been used extensively in vitr...

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“…parahaemolyticus, but the production of delta-VPH by the donor vibrio has not been demonstrated (Taniguchi et al, 1990).…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of a lecithin-dependent haemolysin from Escherichia coli transformed by a Vibrio parahaemolyticus gene

Shinoda

Matsuoka

Tsuchie

et al. 1991

Journal of General Microbiology

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Lecithin-dependent haemolysin (LDH) of Vibrio puruhuemolyticus was purified from Escherichia coli C600 transformed with a plasmid (pHL591) ligated with a 1.5 kb DNA fragment of V. puruhuemolyticus. The final preparation comprised two LDH proteins with different molecular masses which were immunologically crossreactive and had the same enzymic activity. The LDH was a phospholipase hydrolysing both fatty acid esters of phospholipid, i.e. it hydrolysed phosphatidylcholine (PC) to lysophosphatidylcholine (LPC) and then LPC to glycerophosphorylcholine (GPC). From this point of view, LDH should be classified as a phospholipase B.Phospholipase B, however, does not usually show haemolytic activity, because the intermediate (LPC), which is the actual haemolytic agent, is immediately hydrolysed to the final product (GPC). On the other hand, LPC formed by LDH action was comparatively stable, because the rates of the two reactions catalysed by LDH, PC to LPC and LPC to GPC, are almost the same. This is the reason that LDH shows haemolytic activity. Therefore, LDH of V. parahuemolyticus is an atypical phospholipase to be designated as phospholipase A2/lysophospholipase.

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Cloning and characterization of a gene encoding a new thermostable hemolysin from Vibrio parahaemolyticus

Cited by 17 publications

References 13 publications

Haemolysins in Vibrio species

Haemolysins in Vibrio species

Screening for a Single-Chain Variable-Fragment Antibody That Can Effectively Neutralize the Cytotoxicity of the Vibrio parahaemolyticus Thermolabile Hemolysin

Purification and characterization of a lecithin-dependent haemolysin from Escherichia coli transformed by a Vibrio parahaemolyticus gene

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