Stereochemical control of ribosomal peptidyltransferase reaction. Role of amino acid side-chain orientation of acceptor substrate

Publisher Website
Google Scholar
Abstract
The substrate specificity of the acceptor site of peptidyltransferase of Escherichia coli 70S ribosomes was investigated in the fMet-tRNA .cntdot. A-U-G .cntdot. 70S ribosome and AcPhe-tRNA .cntdot. poly(U) .cntdot. 70S ribosome systems by using a series of 2''- and 3''-aminoacyldinucleoside phosphates as acceptors. These chemically synthesized compounds are analogs of the 3'' termini of either 2''(3'')-, 2''-, or 3''-aminoacyl tRNA (AA-tRNA) of the types C-A-aa, C-2''-dA-aa, C-3''-dA-aa, C-3''-dA-3''-NH-aa, and C-2''-dA-2''-NH-aa (aa = Phe, D-Phe, Lys, Leu, Ala, Glu, Pro, Gly, Asp, Met and .alpha.-aminoisobutyryl). The 3''-aminoacyl derivatives of optically active amino acids are much better acceptors of N-formyl-L-methionine (fMet) or N-acetyl-L-phenylalanine (AcPhe) residues than the isomeric 2''-aminoacyl derivatives with affinity constant ratios (Km 2''/3'') > 100. C-A(D-Phe) is a weaker acceptor than the corresponding L derivative C-A-Phe. All glycyl derivatives (C-2''-dA-Gly, C-3''-dA-Gly, C-3''-dA-3''-NH-Gly and C-2''-dA-2''-NH-Gly) are good acceptors of the fMet residue, with ratios (Km 2''/3'') of .apprx. 2. On the basis of these results, a model for the stereochemical control of the peptidyltransferase reaction is proposed. It assigns a major role to the orientation of the amino acid side chain in 2''- or 3''-AA-tRNA. A detailed model of the interaction of the acceptor terminus of 3''-AA-tRNA with the acceptor site of peptidyltransferase is also proposed. The model is strikingly similar to those for the active sites of proteolytic enzymes.