TRANSLOCASE-BOUND SECA IS LARGELY SHIELDED FROM THE PHOSPHOLIPID ACYLCHAINS

Protein translocation in Escherichia coli is mediated by the SecA ATPa se bound to the SecYEG membrane protein complex. SecA translocation AT Pase activity as well as protein translocation is dependent on the pre sence of negatively charged lipids. By using a phospholipid with an ac yl chain linked photoactivatable group, the lipid accessibility of Sec A bound at the translocase was explored. SecA bound to lipid vesicles containing negatively charged lipids was found to be readily accessibl e for labeling by the photoactivatable phospholipid. The presence of a n excess amount of SecYEG complex resulted in a remarkable reduction i n the amount of lipid-accessible SecA irrespective of the nucleotide-b ound form of SecA. These data demonstrate that the SecYEG-bound SecA i s largely shielded from the phospholipid acyl chains and suggest the p resence of two distinct pools of membrane-bound SecA that differ in th e degree of lipid association.