EXPRESSION OF A CLONED CELLULASE XYLANASE GENE FROM PREVOTELLA-RUMINICOLA IN BACTEROIDES-VULGATUS, BACTEROIDES UNIFORMIS AND PREVOTELLA-RUMINICOLA

A new shuttle vector, pRH3 (8.7 kb), was constructed for use in Prevot ella/Bacteroides host strains. This vector combines the pRR12 replicon from P. ruminicola, pBluescript sequences and a tetQ marker gene for selection in Prevotella/Bacteroides hosts. Following insertion of a fr agment carrying an endoglucanase/xylanase gene from P. ruminicola 23 i nto the multiple cloning site, the resulting construct, pRH3X, was int roduced into B. vulgatus 1447, B. uniformis 1100 and P. ruminicola 220 2. This resulted in increases of between 4 and 50-fold in CM-cellulase and xylanase activities in cells grown with glucose. In contrast acti vities were barely detectable for the same construct in E. coli DH5 al pha. Most of the total xylanase activity produced was found within the cell in P. ruminicola 2202 and B. vulgatus 1447 transformed with pRH3 X, and in P. ruminicola 23, An osmotic shock experiment indicated that a significant proportion of the xylanase activity in B. vulgatus 1447 cells carrying pRH3X was periplasmic.