Characterization of an acidic chitinase from seeds of black soybean (Glycine max (L) Merr Tainan No. 3).

Autor: Ya-Min Chang, Li-Chun Chen, Hsin-Yi Wang, Chui-Liang Chiang, Chen-Tien Chang, Yun-Chin Chung
Jazyk: angličtina
Rok vydání: 2014
Předmět:
Zdroj: PLoS ONE, Vol 9, Iss 12, p e113596 (2014)
Druh dokumentu: article
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0113596
Popis: Using 4-methylumbelliferyl-β-D-N,N',N″-triacetylchitotrioside (4-MU-GlcNAc3) as a substrate, an acidic chitinase was purified from seeds of black soybean (Glycine max Tainan no. 3) by ammonium sulfate fractionation and three successive steps of column chromatography. The purified chitinase was a monomeric enzyme with molecular mass of 20.1 kDa and isoelectric point of 4.34. The enzyme catalyzed the hydrolysis of synthetic substrates p-nitrophenyl N-acetyl chitooligosaccharides with chain length from 3 to 5 (GlcNAcn, n = 3-5), and pNp-GlcNAc4 was the most degradable substrate. Using pNp-GlcNAc4 as a substrate, the optimal pH for the enzyme reaction was 4.0; kinetic parameters Km and kcat were 245 µM and 10.31 min-1, respectively. This enzyme also showed activity toward CM-chitin-RBV, a polymer form of chitin, and N-acetyl chitooligosaccharides, an oligomer form of chitin. The smallest oligomer substrate was an N-acetylglucosamine tetramer. These results suggested that this enzyme was an endo-splitting chitinase with short substrate cleavage activity and useful for biotechnological applications, in particular for the production of N-acetyl chitooligosaccharides.
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