Cloning, sequencing, and expression of the cellulase genes of Humicola grisea var. thermoidea.
We have cloned an endoglucanase (EGI) gene and a cellobiohydrolase (CBHI) gene of Humicola grisea var. thermoidea using a portion of the Trichoderma reesei endoglucanase I gene as a probe, and determined their nucleotide sequences. The deduced amino acid sequence of EGI was 435 amino acids in length and the coding region was interrupted by an intron. The EGI lacks a hinge region and a cellulose-binding domain. The deduced amino acid sequence of CBHI was identical to the H. grisea CBHI previously reported, with the exception of three amino acids. The H. grisea EGI and CBHI show 39.8% and 37.7% identity with T. reesei EGI, respectively. In addition to TATA box and CAAT motifs, putative CREA binding sites were observed in the 5' upstream regions of both genes. The cloned cellulase genes were expressed in Aspergillus oryzae and the gene products were purified. The optimal temperatures of CBHI and EGI were 60 degrees C and 55-60 degrees C, respectively. The optimal pHs of these enzymes were 5.0. CBHI and EGI had distinct substrate specificities: CBHI showed high activity toward Avicel, whereas EGI showed high activity toward carboxymethyl cellulose (CMC).
Takashima
S
S
Department of Biotechnology, Faculty of Agriculture, University of Tokyo, Japan.
Nakamura
A
A
Hidaka
M
M
Masaki
H
H
Uozumi
T
T
D000595:Amino Acid Sequence; D001483:Base Sequence; D001709:Biotechnology; D002480:Cellulase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D043366:Cellulose 1,4-beta-Cellobiosidase; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D015870:Gene Expression; D005800:Genes, Fungal; D003904:Mitosporic Fungi / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D017931:DNA Primers; D004271:DNA, Fungal; D002480:Cellulase; D043366:Cellulose 1,4-beta-Cellobiosidase