Abstract
In Escherichia coli, transmembrane translocation of proteins can proceed by a number of routes. A subset of periplasmic proteins are exported via the Tat pathway to which proteins are directed by N-terminal "transfer peptides" bearing the consensus (S/T)RRXFLK "twin-arginine" motif. The Tat system involves the integral membrane proteins TatA, TatB, TatC, and TatE. Of these, TatA, TatB, and TatE are homologues of the Hcf106 component of the DeltapH-dependent protein import system of plant thylakoids. Deletion of the tatB gene alone is sufficient to block the export of seven endogenous Tat substrates, including hydrogenase-2. Complementation analysis indicates that while TatA and TatE are functionally interchangeable, the TatB protein is functionally distinct. This conclusion is supported by the observation that Helicobacter pylori tatA will complement an E. coli tatA mutant, but not a tatB mutant. Analysis of Tat component stability in various tat deletion backgrounds shows that TatC is rapidly degraded in the absence of TatB suggesting that TatC complexes, and is stabilized by, TatB.
Highlights
In Escherichia coli, transmembrane translocation of proteins can proceed by a number of routes
As with the tatB P22L mutant, the tatB::kanR insertion mutant was found to be defective in the export of trimethylamine N-oxide (TMAO) reductase
E. coli TatA and TatE function can be substituted by TatA from Helicobacter pylori, suggesting that only weak amino acid sequence constraints are placed on these essential Tat components
Summary
FϪ, ⌬lacU169,araD139, rpsL150, relA1, pts,F rbsR, flbB5301 MC4100, ⌬tatA ⌬tatE JARV16, pcnB1 zad-981ϻTn10d (KanR) MC4100, ⌬tatB BØD, pcnB1 zad-981ϻTn10d (KanR) MC4100, tatBϻKanR MC4100, ⌬tatABCD, ⌬tatE MC4100, ⌬hyaB⌬hybC MC4100, ⌬hycE. Structed allowing determination of the phenotype of an unambiguously null tatB allele, as well as direct phenotypic comparison with previously characterized in-frame deletion mutants in tatA, tatC, and tatE [10, 12]. The effects of ⌬tatB and tatB::kanR mutations on the localization of an extensive range of Tat-dependent substrates have been assessed. Our studies show that TatB is an essential component of the Tat pathway for all the substrates tested. Complementation analysis is used to support the proposal that TatA and TatE are functionally equivalent to one another, but are functionally distinct from the TatB protein. E. coli TatA and TatE function can be substituted by TatA from Helicobacter pylori, suggesting that only weak amino acid sequence constraints are placed on these essential Tat components. In vivo pulse-chase experiments demonstrate that the TatB protein plays a key role in stabilizing TatC, providing initial evidence for possible complex formation between TatB and TatC
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