In Vivo Experimental Evolution: Re-evolving RNase P RNA

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9981394DoritThis project proposes to re-evolve a critical component of the metabolic machinery of all living cells, the ribonucleoprotein RNaseP. The experimental evolution system that permits replacement of the naturally occurring RNaseP found in the E. coli bacterium with alternate, engineered versions. These synthetic versions, derived either from the variants of RNaseP present in related bacteria, or designed in the laboratory, are placed on a plasmid vector and used to displace the endogenous RNaseP. Repeated generations of in vivo experimental selection result in the evolution of fully functional alternatives that can then be compared and contrasted to their naturally occurring counterparts.This study is one of the first explicit attempts to uncover the existence of alternate functional versions of important metabolic components that are not seen in the natural world. While the versions of RNaseP present in contemporary organism are the exquisite result of billions of years of natural evolution, they likely represent but a small subset of possible solutions to the catalytic challenges facing bacteria. This system delves into these roads not taken, and explores the space separating existing solutions to a given catalytic challenge. In so doing, the relative roles played by contingency, chance, and selection can be understood. This system also provides a general tool with which to probe the functional flexibility of any component of the metabolic machinery, as well as a method for testing the performance of engineered alternatives to naturally occurring proteins and nucleic acids.H:\popbio\abstracts\fy2000\9981394 (Dorit abstract)