Book of Abstracts: Albany 2009
June 16-20 2009
© Adenine Press (2008)
Functional Interactions Between the Ribosome and the Nascent Peptide
Functional interactions between the ribosome and the nascent peptide play an important role in regulation of expression of some bacterial genes. The molecular mechanisms of the nascent peptide recognition are unclear and the extent to which this type of gene regulation is utilized by the cell is unknown. We are interested in identifying the nascent peptide sensors in the ribosome and delineating features of the nascent peptide recognized by these sensors. We investigated programmed drug-dependent ribosome stalling used for regulation of expression of certain antibiotic-resistance genes. Such stalling is controlled by the sequence of the nascent peptide and an antibiotic molecule that binds in the ribosome exit tunnel. We identified the sites of the ribosome stalling and the critical sequences of the nascent peptides required for the erythromycin-dependent stalled complex formation at the regulatory open reading frames of a number of macrolide resistance genes. The results indicate that a variety of nascent peptide sequences can be recognized by the ribosome as stalling signals. We further tested several nucleotides in the ribosome exit tunnel for their potential role in recognition of the nascent peptide. Besides the previously identified nucleotide A2062, a conserved adenine residue at position 2503 appears to play a critical role in sensing the nature of the nascent peptide. Several other nucleotides in the exit tunnel are also involved in either sensing the nascent peptide or forming the stalled translation complex.
In order to determine the extent to which cellular gene regulation is controlled by ribosome-nascent peptide interactions, we compared the proteome of wild type E. coli cells to that of a strain carrying a ribosomal mutation A2058G that affects the nascent peptide recognition. We could obtain conclusive quantitative data from 239 identified proteins. The steady-state levels of at least 13% of the proteins were significantly different (more than 2-fold) between the mutant and wild type. This result suggests that specific interactions between the nascent peptide and the ribosome controls expression of a considerable number of cellular genes. Bioinformatics analysis is currently being carried out to specifically identify cistrons encoding nascent peptides potentially involved in functional interactions with the ribosome.
Center for Pharmaceutical Biotech.