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Characterization of an inhibitor-resistant endo-1,4-β-mannanase from the gut microflora metagenome of Hermetia illucens.
- Source :
- Biotechnology Letters; Oct2018, Vol. 40 Issue 9/10, p1377-1387, 11p
- Publication Year :
- 2018
-
Abstract
- Objective: Hermetia illucens is a voracious insect scavenger that efficiently decomposes food waste. To exploit novel hydrolytic enzymes from this insect, we constructed a fosmid metagenome library using unculturable H. illucens intestinal microorganisms.Results: Functional screening of the library on carboxymethyl cellulose plates identified a fosmid clone with a product displaying hydrolytic activity. Fosmid sequence analysis revealed a novel mannan-degrading gene (ManEM17) composed of 1371 base pairs, encoding 456 amino acids with a deduced 54 amino acid N-terminal signal peptide sequence. Conceptual translation and domain analysis revealed that sequence homology was highest (46%) with endo-1,4-β-mannosidase of Anaerophaga thermohalophila. Phylogenetic and domain analysis indicated that ManEM17 belongs to a novel β-mannanase containing a glycoside hydrolase family 26 domain. The recombinant protein (rManEM17) was expressed in Escherichia coli, exhibiting the highest activity at 55 °C and pH 6.5. The protein hydrolyzed substrates with β-1,4-glycosidic mannoses; maximum specific activity (5467 U mg<superscript>−1</superscript>) occurred toward locust bean gum galactomannan. However, rManEM17 did not hydrolyze p-Nitrophenyl-β-pyranosides, demonstrating endo-form mannanase activity. Furthermore, rManEM17 was highly stable under stringent conditions, including polar organic solvents as well as chemical reducing and denaturing reagents.Conclusions: ManEM17 is an attractive candidate for mannan degradation under the high-organic-solvent and protein-denaturing processes in food and feed industries. [ABSTRACT FROM AUTHOR]
- Subjects :
- METAGENOMICS
GARBAGE disposal units
HOMOLOGY theory
HOMOLOGY (Biochemistry)
ENZYMES
Subjects
Details
- Language :
- English
- ISSN :
- 01415492
- Volume :
- 40
- Issue :
- 9/10
- Database :
- Complementary Index
- Journal :
- Biotechnology Letters
- Publication Type :
- Academic Journal
- Accession number :
- 131779970
- Full Text :
- https://doi.org/10.1007/s10529-018-2596-2