CELA FROM BACILLUS-LAUTUS PL236 ENCODES A NOVEL CELLULOSE-BINDING ENDO-BETA-1,4-GLUCANASE

celA from the cellulolytic bacterium Bacillus lautus PL236 encodes EG-A, an endo-beta-1,4-glucanase. An open reading frame of 2,100 bp preceded by a ribosome-binding site encodes a protein with a molecular mass of 76,863 Da with a typical signal sequence. The NH2-terminal active domain of EG-A is not homologous to any reported cellulase or xylanase and may represent a new family of such enzymes. A 150-amino-acid COOH-terminal peptide is homologous to noncatalytic domains in several other cellulases (A. Meinke, N. R. Gilkes, D. G. Kilburn, R. C. Miller, Jr., and R. A. J. Warren, J. Bacteriol. 173:7126-7135, 1991). Upstream of celA, a partial open reading frame encodes a 145-amino-acid peptide which also belongs to the family mentioned. Zymogram analysis of extracts from Escherichia coli and supernatants of Bacillus subtilis and B. megaterium, including protease-deficient mutants thereof, which express celA, revealed two active proteins, EG-A-L and EG-A-S, with M(r)s of 74,000 and 57,000, respectively. The proportion of EG-A-L to EG-A-S depends on the extracellular proteolytic activity of the host organism, indicating that EG-A-S arises from posttranslational proteolytic modification of EG-A-L. Since EG-A-S has an NH2 terminus corresponding to the predicted NH2-terminal sequence of EG-A, processing appears to take place between the catalytic and noncatalytic domains described. EG-A-L and EG-A-S were purified to homogeneity and shown to have almost identical characteristics with respect to activity against soluble substrates and pH and temperature dependency. EG-A-L binds strongly to cellulose, in contrast to EG-A-S, and has higher activity against insoluble substrates than the latter. We conclude that the COOH-terminal 17,000-M(r) peptide of EG-A-L constitutes a cellulose-binding domain.