ESCHERICHIA-COLI SECB STIMULATES EXPORT WITHOUT MAINTAINING EXPORT COMPETENCE OF RIBOSE-BINDING PROTEIN SIGNAL SEQUENCE MUTANTS

Ribose-binding protein (REP) is exported to the periplasm of Escherich ia coli via the general export pathway. An rbsB-lacZ gene fusion was c onstructed and used to,select mutants defective in REP export. The spo ntaneous Lac(+) mutants isolated in this selection contained either si ngle-amino acid substitutions or a deletion of the REP signal sequence . Intact rbsB genes containing eight different point mutations in the signal sequence were reconstructed, and the effects of the mutations o n REP export were examined. Most of the mutations caused severe defect s in REP export. In addition, different suppressor mutations in SecY/P rlA protein were analyzed for their effects on the export of REP signa l sequence mutants in the presence or absence of SecB. Several REP sig nal sequence mutants were efficiently suppressed, but others were not suppressed. Export of an REP signal sequence mutant in prlA mutant str ains was partially dependent on SecB, which is in contrast to the SecB independence of wild-type REP export. However, the kinetics of export of an REP signal sequence mutant point to a rapid loss of pre-REP exp ort competence, which occurs in strains containing or lacking SecB. Th ese results suggest that SecB does not stabilize the export-competent conformation of REP and may affect translocation by stabilizing the bi nding of pre-REP at the translocation site.