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 β-barrel proteins. The Freeman-Wimley algorithm, which is a highly accurate prediction method based on the physicochemical properties of experimentally characterized transmembrane β 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 β-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 coli; YshA has a β-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 ~0.7nm. Taken together, these data substantiate the predictions made by the Freeman-Wimley algorithm by showing that YshA is a TMBB protein.
Freeman
Thomas C
TC
Department of Biochemistry, Tulane University Health Sciences Center, 1430 Tulane Ave SL-43, New Orleans, LA 70112, USA.
Landry
Samuel J
SJ
Wimley
William C
WC
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism; D016680:Genome, Bacterial; D008051:Lipid Bilayers / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D012486:Salmonella typhimurium / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D008051:Lipid Bilayers; D008081:Liposomes; D011994:Recombinant Proteins; C556258:YshA protein, Salmonella typhimurium