[Interaction of substrates and substrate-like inhibitors with the peptidyltransferase center from Escherichia coli ribosomes].
The binding isotherms of CACCA(3'NHPhe----Ac) and CACCA(3'NHPhe) to E. coli ribosomes and 50S subunits were measured. A theoretical model of adsorption for the case of cooperative interaction between two ligands adsorbed on a ribosome was designated. The analysis of the experimental binding isoterms leads to the following conclusions. A ribosome (or subunit) binds one CACCA (3'NHPhe----Ac) molecule to donor site of the peptidyl transferase center, but two CACCA (3'NHPhe) molecules to both donor and acceptor sites. The binding of CACCA (3'NHPhe) to ribosomes (or subunits) is a cooperative process, characterized by the cooperativity coefficient tau = 40 +/- 5 or more. When model substrates CACCA-Phe, CACCA-Leu and CACCA-Val were taken instead of CACCA (3'NHPhe) in the incubation mixture with ribosomes, dipeptides were obtained even in the case, when ratio [model substrate]: [ribosome] (in moles) was much lower than 1. Puromycin binding to acceptor site with constant (1-2) X 10(4) M-1 also stimulates CACCA(3'NHPhe----Ac) adsorption to the donor site of ribosomes with cooperativity coefficient being equal to 1.5-2.5. It is also shown that cytidine 5'-phosphate binding to the donor site increases kappa cat of the reaction of minimal donors with CACCA-Phe by 1.5 orders of magnitude but has no effect on Km of this reaction. These facts point out that cytidine 5'-phosphate being adsorbed on the corresponding area of the donor site leads to the conversion of low-productive complex [ribosome + minimal donor substrate + acceptor substrate] into high-productive complex [ribosome + minimal donor substrate + acceptor substrate + cytidine 5'-phosphate].
D000217:Acyltransferases / Q000378:metabolism*; D000327:Adsorption; D001665:Binding Sites; D003597:Cytosine Nucleotides / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism*; D007700:Kinetics; D008024:Ligands; D008954:Models, Biological; D010458:Peptidyl Transferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012329:RNA, Bacterial / Q000378:metabolism; D012335:RNA, Ribosomal / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012270:Ribosomes / Q000201:enzymology / Q000378:metabolism*; D013379:Substrate Specificity
D003597:Cytosine Nucleotides; D008024:Ligands; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012346:RNA, Transfer, Amino Acyl; D000217:Acyltransferases; D010458:Peptidyl Transferases
Vzaimodeĭstvie substratov i substratpodobnykh ingibitorov s peptidiltransferaznym tsentrom ribosom Escherichia coli.