Abstract
In Escherichia coli, the Tat system does not translocate Tat signal sequence fused PhoA (RR-PhoA), as it requires disulfide formation for folding. Here we show that such a RR-PhoA construct can be efficiently targeted to the Tat translocon, but the transport is not completed. RR-PhoA is detectable in a 580-kDa TatBC-containing complex, which is the first substrate-bound TatBC complex detected in a bacterial system so far. A second TatBC complex near 440 kDa comprises most of the TatB and TatC but is devoid of RR-PhoA. The targeting of PhoA to the Tat translocon depends on the twin-arginine motif and results in severe growth defects. This physiological effect is likely to be due to proton leakage at the cytoplasmic membrane. The results point to mechanistic incompatibilities of the Tat system with unfolded proteins such as RR-PhoA. There does not exist an intrinsic quality control at the TatBC complex itself, although correct folding is inevitable for Tat-dependent translocation.
Highlights
In E. coli, it has been shown that unfolded proteins are not translocated by the Tat system [12, 13]
We used in this study a Tat signal sequence/PhoA fusion protein to address the question whether there exists a general selection for folded proteins at the Tat translocon
The Tat system does not translocate unfolded proteins, such as apoforms of c-type cytochromes or reduced alkaline phosphatase, when such proteins are N-terminally fused to Tat signal sequences [12, 13]
Summary
In E. coli, it has been shown that unfolded proteins are not translocated by the Tat system [12, 13]. PhoA normally cannot fold properly in the cytoplasm due to a necessity of disulfide formation, and it was observed that Tat signal sequence fused PhoA was not translocated by the Tat system [13]. Tat-dependent translocation could be achieved if cytoplasmic disulfide formation was allowed, a folding prerequisite for PhoA. Based on these results, a general quality control step was proposed [13, 14]. Tat signal sequence binding chaperones are known to optimize proper folding prior to targeting of specific cofactor containing Tat substrates, and this can be important for the formation of an active enzyme, as has been shown for E. coli hydrogenase-2 [15]. Albeit the translocon itself does not reject unfolded Tat substrates such as RR-PhoA, the folded state of Tat substrates is a very important criterion for successful translocation
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
