Quality Control of Protein Translation

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DESCRIPTION: (provided by applicant): Bacteria possess a unique system for rescuing aberrantly stalled ribosomes and marking for degradation the still linked, partially synthesized protein fragments. This quality control system, also known as transtranslation, is orchestrated by a remarkable RNA (SsrA RNA) that functions as a tRNA to detect and revive stalled ribosomes and as an mRNA to facilitate the addition of a short degradation tag to the C-terminus of nascent polypeptides. All known activities of SsrA require SmpB, a small protein that binds SsrA specifically and with high affinity to promote its association with stalled ribosomes. The molecular basis for the formation of the SmpB SsrA complex and the subsequent recognition of impaired ribosomes are not well understood. The objective of this research program is to use a combination of molecular genetics, protein biochemistry, bioinformatics, and structural approaches to elucidate the mechanism of the SmpB-SsrA quality control system. The emphasis is on the molecular characterization of how SmpB recognizes SsrA RNA and promotes the detection and rescue of stalled ribosomes. Principally, through these studies we wish to understand the biochemical and structural basis for the interactions of SmpB with SsrA RNA. Specifically we want to learn what amino acid residues are involved, what base-specific contacts are made, and what structural features contribute to the formation of the SmpB SsrA complex and its interaction with the ribosome. Furthermore, we wish to identify and characterize any additional cellular factors that might participate in this process. Specific complexes of RNA and protein perform many essential biological functions, including RNA processing, RNA turnover, RNA transport, RNA folding, as well as the translation of genetic information from mRNA into protein sequences. Principles that govern RNA-protein interactions are inadequately understood due in large part to a paucity of structural information on RNA-protein complexes. These principles are important for understanding RNA-protein machines, such as the ribosome, and RNA-protein structure and function in general. The relative simplicity of the SmpB-SsrA interaction, the stability of the complex, and recruitment of additional novel factors during trans-translation makes it an ideal system to study the basic principles underlying the assembly of RNA-protein complexes. Understanding of the RNA-protein assembly processes in this system are likely to provide new insights generalizable to the molecular mechanism of how RNA-binding proteins function. Moreover, because the SmpB SsrA quality control system exists only in prokaryotes and involves novel RNA and protein factors that are essential for the survival of most (if not all) pathogenic bacteria, a better understanding of this unique process might allow the design of highly specific new anti-bacterial agents.