Identical RNA-Protein Interactions in Vivo and in Vitro and a Scheme of Folding the Newly Synthesized Proteins by Ribosomes

Debasis Das,Salman Hasan,Chanchal Das Gupta,Pradip Ghorai,Anindita Das,Arpita Bhattacharya,Abhijit Chakrabarti,Santanu Dasgupta,Dibyendu Samanta

doi:10.1074/jbc.m112.396127

Abstract

A distinct three-dimensional shape of rRNA inside the ribosome is required for the peptidyl transfer activity of its peptidyltransferase center (PTC). In contrast, even the in vitro transcribed PTC RNA interacts with unfolded protein(s) at about five sites to let them attain their native states. We found that the same set of conserved nucleotides in the PTC interact identically with nascent and chemically unfolded proteins in vivo and in vitro, respectively. The time course of this interaction, difficult to follow in vivo, was observed in vitro. It suggested nucleation of folding of cytosolic globular proteins vectorially from hydrophilic N to hydrophobic C termini, consistent with our discovery of a regular arrangement of cumulative hydrophobic indices of the peptide segments of cytosolic proteins from N to C termini. Based on this observation, we propose a model here for the nucleation of folding of the nascent protein chain by the PTC.

Highlights

Ribosomal peptidyltransferase center (PTC) acts as a protein folding modulator in vivo and in vitro
We show that a nascent protein interacts with the same set of nucleotides in the PTC in vivo
We have shown that the nascent protein interacts in vivo with the rRNA nucleotides of the PTC identically to how the chemically unfolded protein interacts with the 23 S rRNA or in vitro transcribed PTC RNA

Summary

Background

Ribosomal PTC acts as a protein folding modulator in vivo and in vitro. Results: A fixed set of nucleotides in the PTC interacts to fold polypeptides in vivo and in vitro. The time course of this interaction, difficult to follow in vivo, was observed in vitro It suggested nucleation of folding of cytosolic globular proteins vectorially from hydrophilic N to hydrophobic C termini, consistent with our discovery of a regular arrangement of cumulative hydrophobic indices of the peptide segments of cytosolic proteins from N to C termini. The unfolded forms of a number of unrelated proteins interact with specific sites of the PTC RNA to fold gradually (18 –20) This in vitro activity of PTC prompted us to check whether it has any physiological relevance. The time course of this nucleotide-amino acid interaction cannot be followed in vivo, but we could follow it in vitro It suggests that the nucleation of folding takes place from the N to C terminus of the linear polypeptide

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION