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Collagenolytic subtilisin-like protease from the deep-sea bacterium Alkalimonas collagenimarina AC40T
- Kurata, Atsushi, Uchimura, Kohsuke, Kobayashi, Tohru, Horikoshi, Koki
- Applied microbiology and biotechnology 2010 v.86 no.2 pp. 589-598
- EDTA (chelating agent), Escherichia coli, active sites, amino acids, bacteria, casein, collagen, gelatin, genes, open reading frames, proteinases
- A new alkaline protease (AcpII) was purified from a culture of the deep-sea bacterium Alkalimonas collagenimarina AC40T. AcpII degraded collagen three times faster than it degraded casein. The optimal pH was 8.5-9, and the optimal temperature was 45°C for the degradation of collagen. AcpII was completely inhibited by phenylmethylsulfonyl fluoride and partially by EDTA. Cloning and sequencing the gene for AcpII revealed a 2,283-bp open reading frame encoding a protein of 760 amino acids. AcpII comprises a prepropeptide, a catalytic domain that includes a protease-associated domain (PA domain), and tandem repeat prepeptidase C-terminal domains. To elucidate the role of the PA domain of AcpII, we constructed genes for two enzyme derivatives that possessed the catalytic domains with or without the PA domain and expressed them in Escherichia coli. The derivative without the PA domain showed increased specific activities toward all proteinaceous substrates tested, including gelatin, casein, and collagen, compared with those of the derivative with the PA domain.