SecA alone can promote protein translocation and ion channel activity: SecYEG increases efficiency and signal peptide specificity.

Ying-hsin Hsieh, Hao Zhang, Bor-ruei Lin, Ningren Cui, Bing Na, Hsiuchin Yang, Chun Jiang, Sen-fang Sui, Phang C Tai

The Journal of biological chemistry 2011 Dec 30

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SecA is an essential component of the Sec-dependent protein translocation pathway across cytoplasmic membranes in bacteria. Escherichia coli SecA binds to cytoplasmic membranes at SecYEG high affinity sites and at phospholipid low affinity sites. It has been widely viewed that SecYEG functions as the essential protein-conducting channel through which precursors cross the membranes in bacterial Sec-dependent pathways, and that SecA functions as a motor to hydrolyze ATP in translocating precursors through SecYEG channels. We have now found that SecA alone can promote precursor translocation into phospholiposomes. Moreover, SecA-liposomes elicit ionic currents in Xenopus oocytes. Patch-clamp recordings further show that SecA alone promotes signal peptide- or precursor-dependent single channel activity. These activities were observed with the functional SecA at about 1-2 μM. The results show that SecA alone is sufficient to promote protein translocation into liposomes and to elicit ionic channel activity at the phospholipids low affinity binding sites, thus indicating that SecA is able to form the protein-conducting channels. Even so, such SecA-liposomes are less efficient than those with a full complement of Sec proteins, and lose the signal-peptide proofreading function, resembling the effects of PrlA mutations. Addition of purified SecYEG restores the signal peptide specificity and increases protein translocation and ion channel activities. These data show that SecA can promote protein translocation and ion channel activities both when it is bound to lipids at low affinity sites and when it is bound to SecYEG with high affinity. The latter of the two interactions confers high efficiency and specificity.

Citation

Ying-hsin Hsieh, Hao Zhang, Bor-ruei Lin, Ningren Cui, Bing Na, Hsiuchin Yang, Chun Jiang, Sen-fang Sui, Phang C Tai. SecA alone can promote protein translocation and ion channel activity: SecYEG increases efficiency and signal peptide specificity. The Journal of biological chemistry. 2011 Dec 30;286(52):44702-9