Analysis of Staphylococcus aureus cytoplasmic membrane proteins by isoelectric focusing.
Cytoplasmic membranes were isolated from late-exponential phase Staphylococcus aureus 6539 P and the membrane proteins examined under non-denaturing conditions by thin-layer isoelectric focusing (TLIEF) in a pH 3.5-9.5 gradient. Isolated membrane preparations retained protein integrity as judged by the demonstration of membrane bound adenosine triphosphatase (ATPase) activity in addition to four other solubilized membrane enzyme markers. Membranes were effectively solubilized with 2.5% Triton X-100 (final concentration). Examination of Triton X-100 solubilized membrane preparations established the presence of 22 membrane proteins with isoelectric points between 3.7 and 6.0. The focused proteins displayed the following enzymatic activities and isoelectric points by zymogram methods: ATPase (EC 3.6.1.3), 4.20; malate dehydrogenase (EC 1.1.1.37), 3.90; lactate dehydrogenase (EC 1.1.1.27), 3.85; two membrane proteins exhibited multiple bands upon enzymatic staining NADH dehydrogenase (EC 1.6.99.3), 4.25, 4.35; succinate dehydrogenase (EC 1.3.99.1), 4.85, 5.10, 5.35.
D000251:Adenosine Triphosphatases / Q000032:analysis; D000469:Alkaline Phosphatase / Q000032:analysis; D002462:Cell Membrane / Q000201:enzymology; D007525:Isoelectric Focusing; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D008291:Malate Dehydrogenase / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis*; D009245:NADH Dehydrogenase / Q000032:analysis; D013211:Staphylococcus aureus / Q000201:enzymology*; D013385:Succinate Dehydrogenase / Q000032:analysis
D008565:Membrane Proteins; D007770:L-Lactate Dehydrogenase; D008291:Malate Dehydrogenase; D013385:Succinate Dehydrogenase; D009245:NADH Dehydrogenase; D000469:Alkaline Phosphatase; D000251:Adenosine Triphosphatases