Isolation and Characterization of Extracellular Chitinase Produced by Chitinolytic Bacteria Isolated from Soil Samples
Asian Journal of Microbiology and Biotechnology, Volume 8, Issue 2,
Page 16-24
DOI:
10.56557/ajmab/2023/v8i28225
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.
- Chitinase enzyme
- chitin
- enzyme kinetics
- enzyme activity
How to Cite
References
Zarei M, Aminzadeh S, Ghoroghi A, Motalebi AA, Alikhajeh J, Daliri M. Chitinase isolated from water and soil bacteria in shrimp farming ponds. Iranian J Fish Sci. 2012;11:911–925.
Essghaier B, Hedi A, Bejji M, Jijakli H, Boudabous A, Sadfi-Zouaoui N. Characterization of a novel chitinase from a moderately halophilic bacterium, Virgibacillus marismortui strain M3-23. Ann Microbiol. 2012;62:835–841.
Singh AK. Optimization of culture conditions for thermostable chitinase production by Paenibacillus sp. D1. Afr J Microbiol Res. 2010;4:2291–2298.
Faramarzi MA, Fazeli M, Yazdi MT, Adrangi S, Ahmadi KJA, Tasharrofi N, Mohseni FA. Optimization of cultural conditions for production of chitinase by a soil isolate of Massilia timonae. Biogeosciences. 2009;8:93–99.
Kamil Z, Rizk M, Saleh M, Moustafa S. Isolation and identification of rhizosphere soil chitinolytic bacteria and their potential in antifungal biocontrol. Global J Mol Sci. 2007;2:57–66.
Mubarik NR, Mahagiani I, Anindyaputri A, Santoso S, Rusmana I (2010) Chitinolytic bacteria isolated from chili rhizosphere: chitinase characterization and its application as biocontrol for whitefly (Bemisia tabaci genn.). Am J Agril Biol Sci 5:430–435
Han Y, Li Z, Miao X, Zhang F (2009) Characterization of antifungal chitinase from marine Streptomyces sp. DA11 associated with South China Sea sponge Craniella australiensis. Marine Biotechnol 11:32–40
Budi SW, van Tuinen D, Arnould C, Dumas-Gaudut E, Gianinazzi-Pearson V, Gianinazzi S (2000) Hydrolytic enzyme activity of Paenibacillus sp. strain B2 and effect of antagonistic bacterium on cell wall integrity of two soil-borne pathogenic fungi. Appl Soil Ecol 15:191–199.
Watanable T, Oyanagi W, Suzuki K, Tanaka H. J. Bacteriol. 1990;72:4017-4022.
Patel B, Gohel V, Raol B. Statistical optimization of medium components for chitinase production by Paenibacillus sabinae strain JD2. Ann Microbiol. 2007;57:589–597.
Hui DD, Wei L, Lian WHU, Ying SAX. Effect of medium composition on the synthesis of chitinase and chitin deacetylase from thermophilic Paenibacillus sp. Hu1. Procedia Environ Sci. 2011;8:620–628.
Manucharova NA, Vlasenko AN, Menko EV, Zvyagintsev DG. Specificity of the chitinolytic microbial complex of soils incubated at different temperatures. Microbiol. 2011;80:205–215.
Kuzu SB, Guvenmez HK, Denizci AA. Production of a thermostable and alkaline chitinase by Bacillus thuringiensis subsp. kurstaki strain HBK-51. Biotechnol Res Int. 2012;1–6. DOI:10.1155/2012/135498
Mendonsa ES, Vartak PH, Rao JV, Deshpande MV. An enzyme from Myrothecium verrucavia that degrades insect cuticle from biocontrol of Aedes aegypti mosquito. Biotechnology Letter. 1996;18:373-376.
Harley JP, Prescott LM. Laboratory exercises in microbiology, Fifth edition. The McGraw-Hill Publishing Companies; 2002.
Gram HC. Über die isolierte Färbung der Schizomyceten in Schnitt- und Trockenpräparaten. Fortschritte der Medizin (in German). 1884;2:185–189.
Lokeshwari, Joshi K. Biosorption of Heavy Metal (Chromium) Using Biomass Global Journal of Environmental Research. 2009;3(1):29-35.
Monreal J, Reese ET. The chitinase of Serratia marcescens, Can J Microbiol. 1969;15:689-696.
Lowry OH, Rosebrough J, Farr AL, Randall RJ. Protein measurement with the the folin phenol reagent. Journal of Biological Chemistry. 1951;193:265-275.
Watanabe T, Kobori K, Miyashita K, Fujii T, Sakai H, Uchida M, Tanaka H. Identification of glutamic acid 204 and aspartic acid 200 in chitinase A1 of Bacillus circulans WL-12 as essential residues for chitinase activity. J Biol Chem. 1993;268:18567–72.
Hanson AD, McCarty DR, Henry CS, Xian X, Joshi J, Patterson JA, Fleischmann SD, Tivendale ND, Millar AH. The number of catalytic cycles in an enzyme’s lifetime and why it matters to metabolic engineering. Proceedings of the National Academy of Sciences. 2021;118(13):e2023348118. Available: https://doi.org/10.1073/pnas.2023348118
Felse PA, Panda T. Production of microbial chitinases-a revisit. Bioprocess Eng. 2000;23 Suppl 2:127-34.
Brurberg MB, Synstad B, Klemsdal SS, Aalten DMFV, Sundheim L, Eijsink VGH. Recent Research Development in Microbiology Manuscript; 2000.
Sitrit Y, Vorgias C E, Chet I and Oppenheim AB. Bacteriological Journal. 1995;177(14):4187–4189.
English translation in: Brock TD. Milestones in Microbiology 1546–1940(2 ed.). ASM Press. 1999;215–218.
Andronopoulou E, Vorgias CE. Multiple components and induction mechanism of the chitinolytic system of the hyperthermophilic archaeon Thermococcus chitonophagus. Applied Microbiology and Biotechnology. 2004; 65:694-702.
-
Abstract View: 0 times
PDF Download: 0 times
Download Statistics
Downloads
Current Issue
Subscription
Login to access subscriber-only resources.