Identification of Protein-Protein Interactions Between the TatB and TatC Subunits of the Twin-Arginine Translocase System and the Redox Enzyme Maturation Protein Chaperones

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The Twin-arginine translocation pathway (Tat) serves for targeting and translocation of fully folded proteins across cytoplasmic membrane in bacterial and plant chloroplast thylakoid membranes. TatA, TatB, and TatC are the core components of the Escherichia coli Tat system, where the TatB and TatC subunits are considered to form a receptor complex for Tat-bound proteins. TatB protein is composed of a transmembrane helix and extramembrane part facing the cytoplasmic side of the cell, the structure of TatC revealed six transmembrane helixes and four extramembrane loops. Redox Enzyme Maturation Proteins (REMPs) are the system specific chaperons, which play significant roles in respiratory enzyme maturation, including targeting to the Tat translocase system. We applied a bacterial two-hybrid technique to determine whether REMPs interact with the extramembrane loops of the TatBC recognition module. Individual loops were cloned and fused with one of the two catalytic domains of adenylate cyclase from Bordetella pertussis. The constructs were co-expressed in Escherichia coli where peptides interaction leads to functional complementation between the two adenylate cyclase catalytic domains resulting in increased cAMP levels and induction of beta-galactosidase expression. Our study demonstrated that the DmsD chaperone interacts with the loops 1, 2 and 4 of TatC and with the proposed unfolded domain of TatB. Other E. coli chaperones homologous to DmsD- YcdY and NarJ - also show an interaction with the TatB domain. This suggests that there may be a handoff mechanism of Tat substrates to the translocase via the REMP.