Search National Agricultural Library Digital Collections

NALDC Record Details:

Identification, cloning, and expression of a GHF9 cellulase from Tribolium castaneum (Coleoptera: Tenebrionidae)

Permanent URL:
http://handle.nal.usda.gov/10113/48903
File:
Download [PDF File]
Abstract:
The availability of sequenced insect genomes has allowed for discovery and functional characterization of novel genes and proteins. We report use of the Tribolium castaneum (Herbst) (red flour beetle) genome to identify, clone, express, and characterize a novel endo-β-1,4-glucanase we named TcEG1 (T. castaneum endoglucanase 1). Sequence analysis of a full-length TcEG1 cDNA clone (1356 bp) revealed sequence homology to enzymes in glycosyl hydrolase family 9 (GHF9), and verified presence of a change (Gly for Ser) in the conserved catalytic domain for GHF9 cellulases. This TcEG1 cDNA clone was predicted to encode a 49.5 kDa protein with a calculated pI of 5.39. Heterologous expression of TcEG1 in Drosophila S2 cell cultures resulted in secretion of a 51-kDa protein, as determined by Western blotting. The expressed protein was used to characterize TcEG1 enzymatic activity against two cellulose substrates to determine its specificity and stability. Our data support that TcEG1 as a novel endo-β-1,4-glucanase, the first functional characterization of a cellulase enzyme derived from an insect genome with potential applications in the biofuel industry due to its high relative activity at alkaline pH.
Author(s):
Willis, Jonathan D. , Oppert, Brenda , Oppert, Cris , Klingeman, William E. , Jurat-Fuentes, Juan L.
Subject(s):
Tribolium castaneum , endo-1,4-beta-glucanase , molecular cloning , complementary DNA , sequence analysis , gene expression regulation , enzyme activity
Format:
p. 300-306.
Note:
Includes references
Source:
Journal of insect physiology 2011 Feb., v. 57, issue 2
Language:
English
Year:
2011
Collection:
Journal Articles, USDA Authors, Peer-Reviewed
Rights:
Works produced by employees of the U.S. Government as part of their official duties are not copyrighted within the U.S. The content of this document is not copyrighted.