Analysis of Escherichia coli ribosomal proteins by an improved two dimensional gel electrophoresis. II. Characterization of four new proteins.
Four new proteins, A, B, C, and D, found in Escherichia coli ribosomes by an improved two dimensional gel electrophoresis were characterized by oxidation, reduction, and carboxymethylation of cysteine residues, and CsCl fractionation. The cysteine contents of proteins A, B, C, and D were determined to be 1 +/- 0, 3 +/- 1, 5 +/- 1, and 0 +/- 0 by carboxymethylation with iodoacetic acid. The components of protein complexes, which formed numerously under non-reducing conditions, were analyzed. Including protein A, B, and C, every ribosomal protein (r-protein) having cysteine residue(s) except unconfirmed S1 was proved to form such complexes with various combinations. The cysteine residue in protein A, in particular, was highly reactive to make intermolecular S-S bridges so that spot A almost disappeared on the second dimension gel under the non-reducing conditions. Proteins B and C shifted their spots by reduction towards upper left side as do all known r-proteins having plural cysteine residues except S1. This suggests that proteins B and C change their conformation by intramolecular S-S bridges. The CsCl density gradient centrifugation of high salt washed 70S ribosomes showed that protein A belonged to the insoluble split proteins, proteins B and C to the core particles, and protein D and a small population of B to the soluble split proteins. The electrophoretic behaviors, CsCl fractionation and stoichiometry of the four new proteins suggested strongly that they were intrinsic ribosomal constituents different from known ribosomal proteins or factors.
D001426:Bacterial Proteins / Q000032:analysis* / Q000145:classification; D002499:Centrifugation, Density Gradient; D002586:Cesium; D002712:Chlorides; D003545:Cysteine / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000032:analysis*; D010084:Oxidation-Reduction; D012269:Ribosomal Proteins / Q000032:analysis* / Q000145:classification
D001426:Bacterial Proteins; D002712:Chlorides; D012269:Ribosomal Proteins; D002586:Cesium; C028019:cesium chloride; D003545:Cysteine