An alkaline serine-proteinase from a bacterium isolated from bat feces: purification and characterization.
An alkaline serine-proteinase from Bacillus sp. PN51 isolated from bat feces collected in Phang Nga, Thailand, was purified and characterized. The molecular mass was estimated to be 35.0 kDa. The N-terminal 25 amino acid sequence was about 70% identical with that of Natrialba magadii halolysin-like extracellular serine protease. The enzyme showed the highest proteinase activity at 60 degrees C at pH 10.0. The activity was strongly inhibited by PMSF and chymostatin. The proteinase activity was not affected by the presence of 2% urea, 2% H(2)O(2), 12% SDS, 15% triton X-100, or 15% tween 80. The proteinase preferred Met, Leu, Phe, and Tyr residues at the P(1) position, in descending order. The k(cat), K(m) and k(cat)/K(m) values for Z-Val-Lys-Met-MCA were 16.8+/-0.14 min(-1), 5.1+/-0.28 microM, and 3.3+/-0.28 microM(-1) min(-1) respectively. This is the first report of an alkaline serine-proteinase with extremely high stability against detergents such as SDS.
Tanskul
Somporn
S
Molecular Biotechnology and Bioinformatics, Faculty of Science, Prince of Songkla University, Hat Yai, Thailand. somporn.t@psu.ac.th
Hiraga
Kazumi
K
Takada
Katsumi
K
Rungratchote
Suchart
S
Suntinanalert
Prasert
P
Oda
Kohei
K
D000595:Amino Acid Sequence; D000818:Animals; D001407:Bacillus / Q000166:cytology / Q000201:enzymology*; D002685:Chiroptera / Q000382:microbiology*; D003470:Culture Media; D003902:Detergents / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D005110:Extracellular Space / Q000201:enzymology; D005243:Feces / Q000382:microbiology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000494:pharmacology; D057057:Serine Proteases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature
D003470:Culture Media; D003902:Detergents; D011480:Protease Inhibitors; D057057:Serine Proteases