Characterization and sequence analysis of a Streptomyces rochei A2 endoglucanase-encoding gene.
A 7-kb fragment of Streptomyces rochei A2 chromosomal DNA was cloned into pAT153 and shown to confer endoglucanase (EglS) activity on Escherichia coli cells. In E. coli clones, the EglS was secreted into the periplasm. Deletion analysis revealed that an 827-bp fragment was enough for the enzymatic activity. Sequence analysis showed that the 827-bp fragment codes for the catalytic domain of the enzyme. The complete sequence of the gene (eglS) is 1149-bp long. A signal peptide, a catalytic domain and a cellulose-binding domain were identified from the nucleotide sequence, and the EglS found to belong to the family H of cellulase catalytic domains. These conclusions were substantiated by determination of the N-terminal sequence of the purified protein and zymogram analysis, which revealed protein species with a molecular mass equal to that deduced from the nt sequence analysis.
Perito
B
B
Department of Animal Biology and Genetics Leo Pardi, University of Florence, Italy.
Hanhart
E
E
Irdani
T
T
Iqbal
M
M
McCarthy
A J
AJ
Mastromei
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D002480:Cellulase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012329:RNA, Bacterial / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion / Q000502:physiology; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics*
D012329:RNA, Bacterial; D012333:RNA, Messenger; D002480:Cellulase