PRLA SUPPRESSOR MUTATIONS CLUSTER IN REGIONS CORRESPONDING TO 3 DISTINCT TOPOLOGICAL DOMAINS

The SecY protein of Escherichia coli and its homologues in other organ isms, are integral components of the cellular protein translocation ma chinery. Suppressor mutations that alter SecY (the prlA alleles) broad en the specificity of this machinery and allow secretion of precursor proteins with defective signal sequences. Twenty-five prlA alleles hav e been characterized. These suppressor mutations were found to cluster in regions corresponding to three distinct topological domains of Sec Y. Based on the nature and position of the prlA mutations, we propose that transmembrane domain 7 of SecY functions in signal sequence recog nition. Results suggest that this interaction may involve a right-hand ed supercoil of alpha-helices. Suppressor mutations that alter this do main appear to prevent signal sequence recognition, and this novel mec hanism of suppression suggests a proof-reading function for SecY. We p ropose that suppressor mutations that alter a second domain of SecY, t ransmembrane helix 10, also affect this proof-reading function, but in directly. Based on the synthetic phenotypes exhibited by double mutant s, we propose that these mutations strengthen the interaction with ano ther component of the translocation machinery, SecE. Suppressor mutati ons were also found to cluster in a region corresponding to an amino-t erminal periplasmic domain. Possible explanations for this unexpected finding are discussed.