Abstract
BackgroundMolecular chaperones that support de novo folding of proteins under non stress condition are classified as chaperone ‘foldases’ that are distinct from chaperone’ holdases’ that provide high affinity binding platform for unfolded proteins and prevent their aggregation specifically under stress conditions. Ribosome, the cellular protein synthesis machine can act as a foldase chaperone that can bind unfolded proteins and release them in folding competent state. The peptidyl transferase center (PTC) located in the domain V of the 23S rRNA of Escherichia coli ribosome (bDV RNA) is the chaperoning center of the ribosome. It has been proposed that via specific interactions between the RNA and refolding proteins, the chaperone provides information for the correct folding of unfolded polypeptide chains.ResultsWe demonstrate using Escherichia coli ribosome and variants of its domain V RNA that the ribosome can bind to partially folded intermediates of bovine carbonic anhydrase II (BCAII) and lysozyme and suppress aggregation during their refolding. Using mutants of domain V RNA we demonstrate that the time for which the chaperone retains the bound protein is an important factor in determining its ability to suppress aggregation and/or support reactivation of protein.ConclusionThe ribosome can behave like a ‘holdase’ chaperone and has the ability to bind and hold back partially folded intermediate states of proteins from participating in the aggregation process. Since the ribosome is an essential organelle that is present in large numbers in all living cells, this ability of the ribosome provides an energetically inexpensive way to suppress cellular aggregation. Further, this ability of the ribosome might also be crucial in the context that the ribosome is one of the first chaperones to be encountered by a large nascent polypeptide chains that have a tendency to form partially folded intermediates immediately following their synthesis.
Highlights
Protein folding in biological cells is not yet well understood
The ribosome associated molecular chaperones like the complex of Hsp70 and J-type chaperones in the yeast Saccharomyces cerevisiae and Trigger factor in Escherichia coli ensure that the nascent polypeptide chain is kept in a folding competent state until the whole sequence information is available [1]
Materials Bovine carbonic anhydrase II (BCAII), hen egg white lysozyme, Ribonuclease A, Micrococcus lysodeikticus, cystine hydrochloride, fluorescein isothiocyanate (FITC), Guanidine hydrochloride (GuHCl), dithiothreitol (DTT), GTP, ATP and antibiotics that bind to domainV of 23S rRNA were purchased from Sigma
Summary
Following ribosome mediated synthesis of the proteins the polypeptide chains are released into a highly crowded cellular environment where they require the assistance of a number of molecular chaperones to either fold or be rescued from misfolding and aggregation. The first chaperone to be encountered by these partially folded protein intermediates is likely to be the ribosome since it is itself the site for polypeptide synthesis. The cellular protein synthesis machine can act as a foldase chaperone that can bind unfolded proteins and release them in folding competent state. It has been proposed that via specific interactions between the RNA and refolding proteins, the chaperone provides information for the correct folding of unfolded polypeptide chains
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