3pkp

Publication Abstract from PubMed

Directed evolution is a valuable technique to improve enzyme activity in the absence of a priori structural knowledge, which can be typically enhanced via structure-guided strategies. In this study, a combination of both whole-gene error-prone polymerase chain reaction and site-saturation mutagenesis enabled the rapid identification of mutations that improved RmlA activity toward non-native substrates. These mutations have been shown to improve activities over 10-fold for several targeted substrates, including non-native pyrimidine- and purine-based NTPs as well as non-native d- and l-sugars (both alpha- and beta-isomers). This study highlights the first broadly applicable high throughput sugar-1-phosphate nucleotidyltransferase screen and the first proof of concept for the directed evolution of this enzyme class toward the identification of uniquely permissive RmlA variants.

Expanding the Nucleotide and Sugar 1-Phosphate Promiscuity of Nucleotidyltransferase RmlA via Directed Evolution.,Moretti R, Chang A, Peltier-Pain P, Bingman CA, Phillips GN Jr, Thorson JS J Biol Chem. 2011 Apr 15;286(15):13235-43. Epub 2011 Feb 11. PMID:21317292^[2]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.