Journal
CARBOHYDRATE RESEARCH
Volume 473, Issue -, Pages 46-56Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.carres.2018.12.017
Keywords
Chitinase; Chitin; Melghiribacillus thermohalophilus; Thin-layer chromatography; Hydrolytic pattern
Funding
- Algerian Ministry of Higher Education
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An extracellular acido-thermostable endochitinase (called ChiA-Mt45) from thermohalophilic Melghiribacillus thermohalophilus strain Nari2A(T) gen. nov. sp. nov., was purified and biochemically characterized. The maximum chitinase activity recorded after 48-h of incubation at 55 degrees C was 9000 U/mL. Pure enzyme was obtained after heat treatment (20 min at 90 degrees C) followed by sequential column chromatographies on fast performance liquid chromatography (FPLC) and high performance liquid chromatography (HPLC). Based on MALDI-TOF/MS analysis, the purified enzyme is a monomer with a molecular mass of 45201.10 Da. The 27 residue NH2-terminal sequence of the enzyme showed high homology with Bacillus GH-18 chitinases family. The optimum pH and temperature values for chitinase activity were pH 3.5 and 90 degrees C, respectively. In addition, the enzyme was halotolerant and can be classified as an extremozyme. The pure enzyme was completely inhibited by p-chloromercuribenzoic acid (p-CMB) and N-ethylmaleimide (NEM). Its K-m and k(cat) values were 0.253 mg colloidal chitin/mL and 47000 s(-1), respectively. Interestingly, its catalytic efficiency was higher than those of chitinases ChiA-Hh59 from Hydrogenophilus hirchii KB-DZ44 and chitodextrinase from Streptomyces griseus, and N-acetyl-beta-glucosaminidase from Trichoderma viride. The studied chitinase exhibited high activity towards colloidal chitin, chitin azure, glycol chitin, while it did not hydrolyse chitibiose and amylose. Additionally, thin-layer chromatography (TLC) analysis from chitin-oligosaccharides showed that ChiA-Mt45 acted as an endosplitting enzyme. Overall, the chitinase ChiA-Mt45 may have great potential for the enzymatic degradation of chitin.
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