The Clostridium paraputrificum chiB gene, encoding chitinase B (ChiB), consists of an open reading frame of 2,493 nucleotides and encodes 831 amino acids with a deduced molecular weight of 90,020. The deduced ChiB is a modular enzyme composed of a family 18 catalytic domain responsible for chitinase activity, two reiterated domains of unknown function, and a chitin-binding domain (CBD). The reiterated domains are similar to the repeating units of cadherin proteins but not to fibronectin type III domains, and therefore they are referred to as cadherin-like domains. ChiB was purified from the periplasm fraction of Escherichia coli harboring the chiB gene. The molecular weight of the purified ChiB (87,000) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, was in good agreement with the value (86,578) calculated from the deduced amino acid sequence excluding the signal peptide. ChiB was active toward chitin from crab shells, colloidal chitin, glycol chitin, and 4-methylumbelliferyl -D-N,N-diacetylchitobioside [4-MU-(GlcNAc) 2 ]. The pH and temperature optima of the enzyme were 6.0 and 45°C, respectively. The K m and V max values for 4-MU-(GlcNAc) 2 were estimated to be 6.3 M and 46 mol/min/mg, respectively. SDS-PAGE, zymogram, and Western blot analyses using antiserum raised against purified ChiB suggested that ChiB was one of the major chitinase species in the culture supernatant of C. paraputrificum. Deletion analysis showed clearly that the CBD of ChiB plays an important role in hydrolysis of native chitin but not processed chitin such as colloidal chitin.Chitinase (EC 3.2.1.14) is a glycosyl hydrolase that catalyzes the degradation of chitin, an insoluble linear -1,4-linked polymer of N-acetylglucosamine. Chitinases are present in a wide range of organisms, including bacteria, insects, viruses, plants, and animals, and play important physiological and ecological roles. On the basis of amino acid sequence homology, chitinases are divided into two unrelated families, families 18 and 19 of glycosyl hydrolases (15). Family 18 includes chitinases from bacteria, fungi, viruses, and animals and chitinases classified in class III or V from plants. On the other hand, family 19 includes almost exclusively plant chitinases classified in classes I, II, and IV but also a bacterial chitinase, Streptomyces griseus HUT 6037 chitinase C (32). To date, various chitinases were isolated from aerobic microorganisms such as Bacillus circulans (1,(55)(56)(57), Serratia marcescens (6,13,16,20), an Aeromonas sp. (41,(51)(52)(53), an Alteromonas sp. (48), Streptomyces plicatus (37), Streptomyces olivaceoviridis (3, 36, 38), and Janthiobacterium lividum (12). Several genes encoding chitinases were cloned in Escherichia coli and characterized in detail along with their translated products (9, 12, 20, 36-38, 41, 43, 49, 56, 57). From these studies, chitinases were found to comprise two or more discrete domains, while the function of each domain has not yet been elucidated. Watanabe et al. reported that B. ...