Isolation and Characterization of Extracellular Chitinase Produced by Chitinolytic Bacteria Isolated from Soil Samples

Abstract

Chitin is one of the most common biopolymers found in both marine and terrestrial habitats. Chitinase enzyme has gained popularity due to its diverse biotechnological applications, particularly in agriculture for the biocontrol of phytopathogenic fungi and harmful insects. In this work bacteria producing chitinase enzyme were isolated from soil and water samples. Three bacterial isolates C2, C3, and C4 were found to be Gram-negative coccobacilli while bacterial isolate C9 was found to be Gram-positive cocci. The optimum pH and temperature for all the isolates were studied. The effect of pH, temperature, substrate variation, and incubation period on enzyme activity was studied. The activity of the chitinase enzyme for all the bacterial isolates increased with an increase in temperature up to 37°C. However, a further rise in reaction temperature caused a loss of chitinase activity. Michaelis-Menten plot and Line weaver Burk plot were constructed to calculate the Km and Vmax values. Partial Purification of the chitinase enzyme was carried out and the enzyme activity, protein content, specific activity, and purification fold were estimated. It was observed that the enzyme activity of the crude sample was less than the fractionated sample. The highest enzyme activity was found to be at a 40% fraction. Of the four isolates, C2 was the best in its enzyme activity. The future prospect is the production of chitinases at an industrial scale for various biotechnological applications. Chitinase-producing isolates have the ability to produce chitinase between temperatures 22ºC and 40ºC which is the field temperature for the cultivation of most the crop, so it may be applicable to field conditions against plant pathogenic fungi which is the major problem for agricultural food production. The management of seafood waste businesses may potentially benefit from the usage of this enzyme.