Cloning of the 52-kDa chitinase gene fromSerratia marcescensKCTC2172 and its proteolytic cleavage into an active 35-kDa enzyme
0303 health sciences
572
Base Sequence
Chitinases
Molecular Sequence Data
Enzyme Activation
Isoenzymes
03 medical and health sciences
Bacterial Proteins
Mutagenesis
Endopeptidases
Amino Acid Sequence
Cloning, Molecular
Gene Deletion
Serratia marcescens
DOI:
10.1111/j.1574-6968.1998.tb12905.x
Publication Date:
2006-01-17T09:23:23Z
AUTHORS (8)
ABSTRACT
A chitinase gene (pCHI52) encoding the 52-kDa chitinase was isolated from a Serratia marcescens KCTC2172 cosmid library. This chitinase gene consists of 2526 bp with an open reading frame that encodes 485 amino acids. Escherichia coli harboring the pCHI52 gene secreted not only a 52-kDa but also a 35-kDa chitinase into the culture supernatant. We purified both 52-kDa and 35-kDa chitinases using a chitin affinity column and Sephacryl-S-300 gel filtration chromatography. We determined that the 17 N-terminal amino acid sequences of the 52-kDa and the 35-kDa chitinase are identical. Furthermore, a protease obtained from S. marcescens KCTC2172 cleaved the 52-kDa chitinase into the 35-kDa protein with chitinase activity. These results suggest that the 35-kDa chitinase derives from the 52-kDa chitinase by post-translational proteolytic modification. The optimal reaction temperature of 45 degrees C and the optimal pH of 5.5 were identical for both enzymes. The specific activities of the 52-kDa and 35-kDa chitinases on natural swollen chitin were 67 mumol min-1 mg-1 and 60 mumol min-1 mg-1, respectively.
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