NOVEL CELLULOSE-BINDING DOMAINS, NODB HOMOLOGS AND CONSERVED MODULAR ARCHITECTURE IN XYLANASES FROM THE AEROBIC SOIL BACTERIA PSEUDOMONAS-FLUORESCENS SUBSP CELLULOSA AND CELLVIBRIO-MIXTUS

To test the hypothesis that selective pressure has led to the retentio n of cellulose-binding domains (CBDs) by hemicellulase enzymes from ae robic bacteria, four new xylanase (xyn) genes from two cellulolytic so il bacteria, Pseudomonas fluorescens subsp. cellulosa and Cellvibrio m ixtus, have been isolated and sequenced. Pseudomonas genes xynE and xy nF encoded modular xylanases (XYLE and XYLF) with predicted M(r) value s of 68 600 and 65000 respectively. XYLE contained a glycosyl hydrolas e family 11 catalytic domain at its N-terminus, followed by three othe r domains; the second of these exhibited sequence identity with NodB f rom rhizobia. The C-terminal domain (40 residues) exhibited significan t sequence identity with a non-catalytic domain of previously unknown function, conserved in all the cellulases and one of the hemicellulase s previously characterized from the pseudomonad, and was shown to func tion as a CBD when fused to the reporter protein glutathione-S-transfe rase. XYLF contained a C-terminal glycosyl hydrolase family 10 catalyt ic domain and a novel CBD at its N-terminus. C. mixtus genes xynA and xynB exhibited substantial sequence identity with xynE and xynF respec tively, and encoded modular xylanases with the same molecular architec ture and, by inference, the same functional properties. In the absence of extensive crosshybridization between other multiple eel (cellulase ) and xyn genes from P. fluorescens subsp. cellulosa and genomic DNA f rom C. mixtus, similarity between the two pairs of xylanases may indic ate a recent transfer of genes between the two bacteria.