title: pH-sensitivity of the ribosomal peptidyl transfer reaction dependent on the identity of the A-site aminoacyl-tRNA.
abstract: We studied the pH-dependence of ribosome catalyzed peptidyl transfer from fMet-tRNA(fMet) to the aa-tRNAs Phe-tRNA(Phe), Ala-tRNA(Ala), Gly-tRNA(Gly), Pro-tRNA(Pro), Asn-tRNA(Asn), and Ile-tRNA(Ile), selected to cover a large range of intrinsic pK(a)-values for the α-amino group of their amino acids. The peptidyl transfer rates were different at pH 7.5 and displayed different pH-dependence, quantified as the pH-value, pK(a)(obs), at which the rate was half maximal. The pK(a)(obs)-values were downshifted relative to the intrinsic pK(a)-value of aa-tRNAs in bulk solution. Gly-tRNA(Gly) had the smallest downshift, while Ile-tRNA(Ile) and Ala-tRNA(Ala) had the largest downshifts. These downshifts correlate strongly with molecular dynamics (MD) estimates of the downshifts in pK(a)-values of these aa-tRNAs upon A-site binding. Our data show the chemistry of peptide bond formation to be rate limiting for peptidyl transfer at pH 7.5 in the Gly and Pro cases and indicate rate limiting chemistry for all six aa-tRNAs.
authors.affiliation: Department of Cell and Molecular Biology, Biomedical Center, Uppsala University, Box 596, S-751 24 Uppsala, Sweden.
authors.forename: Magnus
authors.initials: M
authors.lastname: Johansson
authors.forename: Ka-Weng
authors.initials: KW
authors.lastname: Ieong
authors.forename: Stefan
authors.initials: S
authors.lastname: Trobro
authors.forename: Peter
authors.initials: P
authors.lastname: Strazewski
authors.forename: Johan
authors.initials: J
authors.lastname: Åqvist
authors.forename: Michael Y
authors.initials: MY
authors.lastname: Pavlov
authors.forename: Måns
authors.initials: M
authors.lastname: Ehrenberg
mesh_terms: D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008954:Models, Biological; D056004:Molecular Dynamics Simulation; D015394:Molecular Structure; D010455:Peptides / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism*; D012358:RNA, Transfer, Met / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism*
chemical_list: D010455:Peptides; D012333:RNA, Messenger; D012346:RNA, Transfer, Amino Acyl; D012358:RNA, Transfer, Met; C089238:fMet-tRNA(fMet)