Purification and characterization of a cysteine protease produced by pathogenic luminous Vibrio harveyi.
The purification and characterization of an extracellular protease produced by pathogenic luminous Vibrio harveyi strain 820514, originally isolated from diseased tiger prawn (Penaeus monodon), was presented in this paper. The purification steps included ammonium sulfate precipitation, with columns of hydrophobic interaction chromatography and anion exchange on fast protein liquid chromatography. The protease is an alkaline cysteine protease, heat labile, inhibited by iodoacetamide, iodoacetic acid, N-ethylmaleimide, p-chloromercuribenzoate, and p-chloromercuribenzene-sulfonic acid, and showed maximal activities at pH 8 and 50 degrees C, having a molecular mass of 38 kDa as estimated by SDS-PAGE and gel filtration column. In addition, the protease was also completely inhibited by CuCl2 and HgCl2, but not or only partially inhibited by other inhibitors tested. Furthermore, 2-mercaptoethanol was the most effective reducing agent in the activation of the enzyme. The present protease is the first cysteine protease found in Vibrio species.
Liu
P C
PC
Department of Aquaculture, National Taiwan Ocean University, 2, Pei-Ning Road, Keelung, Taiwan 202, ROC.
Lee
K K
KK
Tu
C C
CC
Chen
S N
SN
D003546:Cysteine Endopeptidases / Q000302:isolation & purification*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D013696:Temperature; D014733:Vibrio / Q000201:enzymology*
D015853:Cysteine Proteinase Inhibitors; D003546:Cysteine Endopeptidases