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Applied and Environmental Microbiology, January 2002, p. 263-270, Vol. 68, No. 1
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.1.263-270.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Identification and Characterization of the Gene Cluster Involved in Chitin Degradation in a Marine Bacterium, Alteromonas sp. Strain O-7

Hiroshi Tsujibo,* Hideyuki Orikoshi, Nao Baba, Masahiro Miyahara, Katsushiro Miyamoto, Masahide Yasuda, and Yoshihiko Inamori

Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094, Japan

Received 16 July 2001/ Accepted 16 October 2001

Alteromonas sp. strain O-7 secretes chitinase A (ChiA), chitinase B (ChiB), and chitinase C (ChiC) in the presence of chitin. A gene cluster involved in the chitinolytic system of the strain was cloned and sequenced upstream of and including the chiA gene. The gene cluster consisted of three different open reading frames organized in the order chiD, cbp1, and chiA. The chiD, cbp1, and chiA genes were closely linked and transcribed in the same direction. Sequence analysis indicated that Cbp1 (475 amino acids) was a chitin-binding protein composed of two discrete functional regions. ChiD (1,037 amino acids) showed sequence similarity to bacterial chitinases classified into family 18 of glycosyl hydrolases. The cbp1 and chiD genes were expressed in Escherichia coli, and the recombinant proteins were purified to homogeneity. The highest binding activities of Cbp1 and ChiD were observed when {alpha}-chitin was used as a substrate. Cbp1 and ChiD possessed a chitin-binding domain (ChtBD) belonging to ChtBD type 3. ChiD rapidly hydrolyzed chitin oligosaccharides in sizes from trimers to hexamers, but not chitin. However, after prolonged incubation with large amounts of ChiD, the enzyme produced a small amount of (GlcNAc)2 from chitin. The optimum temperature and pH of ChiD were 50°C and 7.0, respectively.

* Corresponding author. Mailing address: Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094, Japan. Phone and fax: (81-726) 90-1057. E-mail: tsujibo{at}gly.oups.ac.jp.

Applied and Environmental Microbiology, January 2002, p. 263-270, Vol. 68, No. 1
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.1.263-270.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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