The use of a cleavable crosslinking reagent to identify neighboring proteins in the 30-S ribosomal subunit of Escherichia coli.
A cleavable bifunctional reagent, dimethyl 3,3'-dithiobispropionimidate, has been used to crosslink proteins that occupy neighboring positions in the 30-S ribosomal subunit of Escherichia coli. The crosslinked proteins were identified, fully or partly, by their positions in two two-dimensional gel electrophoretic systems, one diagonal and the other quasi-diagonal, in which the complexes were cleaved after the first-dimensional run. It was found to be necessary to block the protein sulfhydryl groups in order to prevent artifactual disulfide crosslinking after extraction of the protein from ribosome. Eleven crosslinked complexes were detected. Four were fully identified: the triplet S4-S5-S8, and the pairs S2-S3, S4-S5, and S5-S8. In five others one component was identified unambiguously. No additional complexes were seen when the longer homologous butyro and capro reagents were used.
D001665:Binding Sites; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000648:ultrastructure*; D007094:Imides; D046911:Macromolecular Substances; D011485:Protein Binding; D011487:Protein Conformation; D012269:Ribosomal Proteins / Q000032:analysis*; D012270:Ribosomes / Q000648:ultrastructure*; D013438:Sulfhydryl Compounds
D004220:Disulfides; D007094:Imides; D046911:Macromolecular Substances; D012269:Ribosomal Proteins; D013438:Sulfhydryl Compounds