Cloning, sequencing and analysis of the ggh-A gene encoding a 1,4-β- d-glucan glucohydrolase from Microbispora bispora

• Published in: Gene 1996-06, Vol.172 (1), p.93-98
• Main Authors: Goyal, Anil K., Eveleigh, Douglas E.
• Format: Article
• Language: English
• Subjects: Actinomycetales - enzymology; Actinomycetales - genetics; actinomycete; Amino Acid Sequence; Base Sequence; beta-Glucosidase - genetics; beta-Glucosidase - metabolism; Binding Sites; Cellulase; cellulolytic; Cellulose - metabolism; Cloning, Molecular; DNA, Recombinant; domains; Glucan 1,4-beta-Glucosidase; glycosyl hydrolase family 3; Molecular Sequence Data; Open Reading Frames; Sequence Homology, Amino Acid; β-glucosidase
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/0378-1119(96)00076-5

The ggh-A gene, encoding a 1,4-β- d-glucan glucohydrolase/β-glucosidase, of Microbispora bispora (Mb) was subcloned and expressed from a 4.0-kb XhoI DNA fragment. The nucleotide sequence of this fragment was determined. Analysis of the sequence revealed one open reading frame (ORF) which encodes a 986-amino-acid (aa) protein with a calculated molecular weight of 107 510. The ggh-A ORF has features typical of an actinomycete gene including high GC content (70.5%) and corresponding biased codon usage. Comparison of the aa sequence of the Mb 1,4-β- d-glucan glucohydrolase (Mbggh-A) with other glycosidases reveals high overall homology to several β-glucosidases and a 1,4-β- d-glucan glucohydrolase belonging to the glycosyl hydrolase family 3. The aa sequence alignments of Mbggh-A and β-glucosidases show that the active site region potentially involves two Asp residues. The aa sequence homology studies revealed a potential two-domain structure for Mbggh-A and other β-glucosidases. Furthermore, Mbggh-A has localized homology to a cellulose-binding domain present in some xylanases. This report is significant, as, to date, 1,4-β- d-glucan glucohydrolases have rarely been reported, though they are assumed to have a critical role in cellulolysis.