Expression and characterization of a metalloprotease from a Vibrio parahaemolyticus isolate.
The extracellular zinc metalloprotease from Vibrio parahaemolyticus (VPM) is a putative virulence factor for host infection. It is synthesized from the vpm gene of V. parahaemolyticus as a polypeptide of 814 amino acids with an estimated molecular mass of 89,833 Da, containing a zinc metalloprotease HEXXH consensus motif. To investigate the enzymatic properties of V. parahaemolyticus metalloprotease, the mature vpm gene was overexpressed in Escherichia coli, and the recombinant protein (rVPM) was purified by a His-binding metal affinity column (>95% purity). The activity of the recombinant protease produced in E. coli was examined by gelatin activity staining and proteolytic activity assays using gelatin and azocasein as substrates. rVPM showed maximum activity at about 37 degrees C and pH 8. The cytotoxicity against flounder gill cells and fish pathogenicity indicated a potential role in pathogenesis.
Luan
Xiaoyan
X
Department of Marine Biology, College of Marine Life Science, Ocean University of China, 5 Yushan Road, Qingdao 266003, Peoples Republic of China.
Chen
Jixiang
J
Zhang
Xiao-Hua
XH
Li
Yun
Y
Hu
Guobin
G
D000818:Animals; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005393:Fish Diseases / Q000382:microbiology / Q000503:physiopathology; D005432:Flounder; D005759:Gastroenteritis / Q000382:microbiology / Q000662:veterinary; D005780:Gelatin / Q000378:metabolism; D005880:Gills / Q000166:cytology; D006801:Humans; D045726:Metalloproteases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D013194:Staining and Labeling / Q000379:methods; D014735:Vibrio Infections / Q000382:microbiology / Q000662:veterinary; D014736:Vibrio parahaemolyticus / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification / Q000472:pathogenicity*; D015027:Zebrafish / Q000382:microbiology
D005780:Gelatin; D045726:Metalloproteases