Biosynthesis and characterization of a thermostable, alkali-tolerant chitinase from Bacillus pumilus JUBCH08 displaying antagonism against phytopathogenic Fusarium oxysporum.

ABSTRACT

The present investigation highlights the process parameters influencing the submerged fermentation of chitinase by Bacillus pumilus JUBCH08, purification and characterization of the enzyme and determination of antagonistic activity of the bacterium against Fusarium oxysporum. Medium supplemented with 0.5 % chitin and peptone, at initial pH 8.0, when incubated at 35 °C for 72 h favored highest chitinase production. The enzyme was purified 25.1-fold to homogeneity. The chitinase was found to be thermostable and alkali-tolerant with maximum activity at pH 8.0 and 70 °C for 1 h. The molecular weight of chitinase was found to be 64 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Mg2+, Co2+, Ca2+ and Mn2+ improved the chitinase activity. The K m and V max values of the enzyme were 0.13 mg/ml and 38.23 U/ml, respectively. When subjected to dual plate assay, the bacterium showed 45 % antagonism against F. oxysporum. Thus, it could be inferred that cultural conditions strongly affected the chitinase production by B. pumilus JUBCH08. The enzyme being thermostable and best functional under alkaline conditions could be useful for the feed industry and related biotechnological applications. Inhibition of F. oxysporum by the culture through lytic mechanism indicates its potentiality as a biocontrol agent.