Structural Insight into a Membrane Intrinsic Acyltransferase from Chlorobium tepidum.

The Lipid A component of the outer membrane of Gram-negative bacteria is an integral part of the permeability barrier known as LPS, which actively prevents the uptake of bactericidal compounds. It is clinically very significant, as it is known to elicit a strong immune response in the humans, through the TLR4 complex. The Lipid A species are synthesized through a highly conserved multistep biosynthetic pathway. The final step is catalyzed by acyltransferases of the HtrB/MsbB family, which are members of a superfamily of enzymes, present in all domains of life with important roles to play in various biological processes. The investigation of a putative dual functioning enzyme which can add both laurate and myristate residues to the (Kdo)2-lipid IVA (precursor of Lipid A) would give a snapshot into the versatility of substrates that these enzymes catalyze. In this study we have cloned and purified to homogeneity, such a putative dual functional acyltransferase from Chlorobium tepidum, and attempted to study the enzyme in more details in terms of its sequence and structural aspects, as it lacks conserved residues with other enzymes of the same family.

Analysis of haloarchaeal twin-arginine translocase pathway reveals the diversity of the machineries.

The twin-arginine translocase (Tat) pathway transports folded proteins across the plasma membrane and plays a critical role in protein transport in haloarchaea. Computational analysis and previous experimental evidence suggested that the Tat pathway transports almost the entire secretome in haloarchaea. The TatC, receptor component of this pathway shows greater variation in membrane topology in haloarchaea than in other organisms. The presence of a unique fourteen-transmembrane TatC homolog (TatCt) in haloarchaea, over and above the expected TatC topological variants, indicates a strong correlation between the additional homologs and the large number of substrates transported via the haloarchaeal Tat pathway. Various combinations of TatC homologs with different topologies-TatCo, TatCt, TatCn, and TatCx have been observed in haloarchaea. In this report, on the basis of these combinations we have segregated all haloarchaeal Tat substrates into two groups. The first group consists of substrates that are transported by TatCt alone, whereas the second group consists of substrates that are transported by the other TatC homologs (TatCo, TatCn, and TatCx). The various haloarchaea TatA components also shows the possible segregation towards the substrates. We have also identified the possible homologs for Tat substrate chaperones, which act as a quality-control mechanism for proper protein folding. Further sequence analysis implies that the two TatC domains of TatCt complement each other's functionally. Substrate analysis also revealed subtle differences between the substrates being transported by various homologs: further experimental analysis is therefore required for better understanding of the complexities of the haloarchaeal Tat pathway.