Both ends of Escherichia coli ribosomal protein S13 are immunochemically accessible in situ.
To investigate the structure of Escherichia coli ribosomal protein S13 in 30S ribosomal subunits, we have previously generated 22 S13 specific monoclonal antibodies and mapped their specific epitopes to the S13 sequence. The availability of these S13 epitopes in situ has been further examined by incubating these monoclonal antibodies with 30S ribosomal subunits and analyzing formation of monoclonal antibody-linked ribosome dimers by sucrose gradients centrifugation. We have found that none of the 22 monoclonal antibodies makes ribosome dimers individually as do typical antisera. However, one monoclonal antibody, designated AS13-MAb 2, reacts with 30S ribosomal subunits to form immunocomplexes sedimenting faster than subunit monomers. When AS13-MAb 2 is paired with any one of three monoclonal antibodies directed to the S13 C-terminal epitopes, dimer formation is observed. Other pairs of monoclonal antibodies directed to distinct S13 epitopes have been tested similarly for dimer formation. Monoclonal antibody AS13-MAb 22, directed to the N-terminal region of 22 residues, also causes subunits to form typical dimers, but only if paired with one of the three monoclonal antibodies directed to the S13 C-terminal region. The close proximity of the epitopes recognized by AS13-MAbs 2 and 22 has been established by the mutual competition between the antibodies binding to intact 30S subunits. These results corroborate our previous observation, using polyclonal antibodies, that S13 has more than one epitope exposed on 30S subunits. Our finding that sequences on both ends of the S13 molecule are immunochemically accessible provides information about the molecular organization of S13 in situ.
Syu
W J
WJ
Department of Biomolecular Chemistry, University of Wisconsin, Madison 53706.
Kahan
L
L
D000911:Antibodies, Monoclonal; D000939:Epitopes; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D007120:Immunochemistry; D012269:Ribosomal Proteins / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D029968:Escherichia coli Proteins; D012269:Ribosomal Proteins; C512838:rpsM protein, E coli