TRANSFER OF GENES FROM PSEUDOMONAS-SACCHAROPHILA TO CONSTRUCT XYLOSE-UTILIZING STRAINS OF ALCALIGENES-EUTROPHUS

About 1500 hybrid broad-host-range plasmids from a genomic library of Pseudomonas saccharophila were individually transferred by conjugation from Escherichia coli to Alcaligenes eutrophus. Direct selection for pentose-utilizing transconjugants yielded three clones capable of grow th on xylose. Growth of P. saccharophila as well as the transconjugant s of A. eutrophus on xylose was relatively slow, exhibiting doubling t imes of about 9.5 h. Plasmid pGN3 harbored by one transconjugant conta ined a 28-kb DNA insert, 16.4 kb of which comprised the minimal inform ation required for xylose utilization by A. eutrophus. At least the xy l genes encoding xylose isomerase and xylulokinase were located within this region, as indicated by their induction during growth of A. eutr ophus (pGN3) on xylose. Southern hybridizations with a heterologous ge ne probe confirmed the presence of these xyl genes. In both P. sacchar ophila and A. eutrophus (pGN3), low activities of several enzymes oper ating in the pentose phosphate and Entner-Doudoroff pathways might lim it the rate of xylose catabolism.