Loading…

Characterization of an inhibitor-resistant endo-1,4-β-mannanase from the gut microflora metagenome of Hermetia illucens

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 o...

Full description

Saved in:
Bibliographic Details
Published in:Biotechnology letters 2018-10, Vol.40 (9-10), p.1377-1387
Main Authors: Song, Jaeeun, Kim, Su-Yeon, Kim, Dae-Hyuk, Lee, Young-Seok, Sim, Joon-Soo, Hahn, Bum-Soo, Lee, Chang-Muk
Format: Article
Language:English
Subjects:
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary: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 −1 ) 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.
ISSN:0141-5492
1573-6776
DOI:10.1007/s10529-018-2596-2