Characteristics of an exochitinase from Streptomyces olivaceoviridis, its corresponding gene, putative protein domains and relationship to other chitinases.
Streptomyces olivaceoviridis efficiently degrades chitin. Shotgun cloning of partially Sau3A-cleaved DNA using the multicopy vector pIJ702 and Streptomyces lividans 66 as host resulted in the identification of the plasmid pCHI O1 which harbours an insert of 4.6 kb. In the presence of chitin as sole carbon source, transformants of S. lividans 66 carrying pCHI O1 or its derivatives with smaller inserts overproduced an exochitinase which was purified to homogeneity. The chitin-inducible enzyme with an isoelectric point of 4.0 shows optimal activity at pH 7.3 and 55 degrees C, has an apparent molecular mass of 47 kDa and is competitively inhibited by the pseudosugar allosamidin. The enzyme was identified as an exochitinase since it generates exclusively chitobiose from chitotetraose, chitohexaose, and colloidal high-molecular mass chitin. Sequence analysis of a reading frame of 1794 base pairs and comparison of the deduced amino-acid sequence allowed the identification of the putative catalytic domain, one region with significant similarity to the type-III module of fibronectin and one domain of unknown function. Multiple sequence alignment and hydrophobic-cluster analysis of 25 chitinolytic enzymes from bacteria, fungi and plants allowed the identification of their characteristic domains. The exochitinase from S. olivaceoviridis shares highest similarity with the chitinase D from Bacillus circulans.
Blaak
H
H
FB Biologie/Chemie, University of Osnabrück, Germany.
Schnellmann
J
J
Walter
S
S
Henrissat
B
B
Schrempf
H
H
D000117:Acetylglucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D000595:Amino Acid Sequence; D001483:Base Sequence; D002688:Chitinases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics*; D014312:Trisaccharides / Q000494:pharmacology
D011994:Recombinant Proteins; D014312:Trisaccharides; C054851:allosamidin; D002688:Chitinases; D000117:Acetylglucosamine