1MP5

Y177F VARIANT OF S. ENTERICA RmlA

Summary for 1MP5

• Entry DOI: 10.2210/pdb1mp5/pdb
• Related: 1IIM 1IIN 1MP3 1MP4
• Descriptor: Y177F VARIANT OF S. ENTERICA RmlA BOUND TO UDP-GLUCOSE, URIDINE-5'-DIPHOSPHATE-GLUCOSE (2 entities in total)
• Functional Keywords: transferase
• Biological source: Salmonella enterica
• Total number of polymer chains: 4
• Total formula weight: 131909.61

Authors

Barton, W.A.,Biggins, J.B.,Jiang, J.,Thorson, J.S.,Nikolov, D.B. (deposition date: 2002-09-11, release date: 2002-10-09, Last modification date: 2024-02-14)

Primary citation

Barton, W.A.,Biggins, J.B.,Jiang, J.,Thorson, J.S.,Nikolov, D.B.
Expanding pyrimidine diphosphosugar libraries via structure-based nucleotidylyltransferase engineering
Proc.Natl.Acad.Sci.USA, 99:13397-13402, 2002

PubMed Abstract: In vitro "glycorandomization" is a chemoenzymatic approach for generating diverse libraries of glycosylated biomolecules based on natural product scaffolds. This technology makes use of engineered variants of specific enzymes affecting metabolite glycosylation, particularly nucleotidylyltransferases and glycosyltransferases. To expand the repertoire of UDP/dTDP sugars readily available for glycorandomization, we now report a structure-based engineering approach to increase the diversity of alpha-d-hexopyranosyl phosphates accepted by Salmonella enterica LT2 alpha-d-glucopyranosyl phosphate thymidylyltransferase (E(p)). This article highlights the design rationale, determined substrate specificity, and structural elucidation of three "designed" mutations, illustrating both the success and unexpected outcomes from this type of approach. In addition, a single amino acid substitution in the substrate-binding pocket (L89T) was found to significantly increase the set of alpha-d-hexopyranosyl phosphates accepted by E(p) to include alpha-d-allo-, alpha-d-altro-, and alpha-d-talopyranosyl phosphate. In aggregate, our results provide valuable blueprints for altering nucleotidylyltransferase specificity by design, which is the first step toward in vitro glycorandomization.

PubMed: 12374866
DOI: 10.1073/pnas.192468299

Experimental method

X-RAY DIFFRACTION (2.75 Å)