The prediction and characterization of YshA, an unknown outer-membrane protein from Salmonella typhimurium

We have developed an effective pathway for the prediction and characterization of novel transmembrane beta-barrel proteins. The Freeman-Wimley algorithm, which is a highly accurate prediction method based on the physicochemical properties of experimentally characterized transmembrane beta barrel (TMBB) structures, was used to predict TMBBs in the genome of Salmonella typhimurium LT2. The previously uncharacterized product of gene yshA was tested as a model for validating the algorithm. YshA is a highly conserved 230-residue protein that is predicted to have 10 transmembrane beta-strands and an N-terminal signal sequence. All of the physicochemical and spectroscopic properties exhibited by YshA are consistent with the prediction that it is a TMBB. Specifically, recombinant YshA localizes to the outer membrane when expressed in Escherichia colt; YshA has a beta-sheet-rich secondary structure with stable tertiary contacts in the presence of detergent micelles or when reconstituted into a lipid bilayer. When in a lipid bilayer, YshA forms a membrane-spanning pore with an effective radius of similar to 0.7 nm. Taken together, these data substantiate the predictions made by the Freeman-Wimley algorithm by showing that YshA is a TMBB protein. (C) 2010 Elsevier B.V. All rights reserved.