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PURIFICATION AND CHARACTERIZATION OF A CHITINASE FROM PEANUT ( ARACHIS HYPOGAEA L.)
Author(s) -
WANG SHAOYUN,
SHAO BIAO,
YE XIUYUN,
RAO PINGFRAN
Publication year - 2008
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/j.1745-4514.2007.00144.x
Subject(s) - chitinase , arachis hypogaea , isoelectric point , affinity chromatography , gel electrophoresis , isoelectric focusing , polyacrylamide gel electrophoresis , biochemistry , sodium dodecyl sulfate , biology , chromatography , ammonium sulfate precipitation , chemistry , enzyme , botany , size exclusion chromatography
A chitinase was isolated from peanut ( Arachis hypogaea L.) seeds. The procedure entailed extraction, ammonium sulfate precipitation, affinity chromatography on Affi‐gel blue gel, and high‐performance liquid chromatography on POROS 20 HQ. There was a 133‐fold increase in specific activity of purified chitinase compared with that of the crude extract. The protein exhibited a molecular mass of 34.4 kDa in sodium dodecyl sulfate‐polyacrylamide gel electrophoresis both under reducing and nonreducing conditions, indicating that it is a monomeric protein. The isoelectric point was 5.1 by isoelectric focusing electrophoresis. Optimal pH activity was 5.4 and optimal temperature was 40–50C. The enzyme was stable below 55C, but was rapidly inactivated when incubated at temperatures above 60C. These results demonstrated that the purified protein was a kind of relatively thermostable chitinase from the peanut seeds.PRACTICAL APPLICATIONS This paper describes the isolation and characterization of a chitinase from Arachis hypogaea L. seeds. Chitinases are listed as the class of pathogenesis‐related proteins, and they have become a popular research field because of their resistance to plant disease. The research we are presenting should help in understanding chitinase functions in plant defense mechanisms. A relatively thermostable enzyme newly obtained, which was stable below 55C, would certainly be significant for its utilization both in enzymology and in plant defense. In particular, a relatively heat‐resistant enzyme from A. hypogaea L. seeds would certainly be helpful for elaborating strategies for constitutive expression of chitinase genes and their manipulation and general utilization in plant defense mechanisms through more recent techniques of genetic manipulation.