Polynucleotide phosphorylase activity may be modulated by metabolites in Escherichia coli

J Biol Chem. 2011 Apr 22;286(16):14315-23. doi: 10.1074/jbc.M110.200741. Epub 2011 Feb 14.

Abstract

RNA turnover is an essential element of cellular homeostasis and response to environmental change. Whether the ribonucleases that mediate RNA turnover can respond to cellular metabolic status is an unresolved question. Here we present evidence that the Krebs cycle metabolite citrate affects the activity of Escherichia coli polynucleotide phosphorylase (PNPase) and, conversely, that cellular metabolism is affected widely by PNPase activity. An E. coli strain that requires PNPase for viability has suppressed growth in the presence of increased citrate concentration. Transcriptome analysis reveals a PNPase-mediated response to citrate, and PNPase deletion broadly impacts on the metabolome. In vitro, citrate directly binds and modulates PNPase activity, as predicted by crystallographic data. Binding of metal-chelated citrate in the active site at physiological concentrations appears to inhibit enzyme activity. However, metal-free citrate is bound at a vestigial active site, where it stimulates PNPase activity. Mutagenesis data confirmed a potential role of this vestigial site as an allosteric binding pocket that recognizes metal-free citrate. Collectively, these findings suggest that RNA degradative pathways communicate with central metabolism. This communication appears to be part of a feedback network that may contribute to global regulation of metabolism and cellular energy efficiency.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Site
  • Citric Acid / chemistry
  • Cloning, Molecular
  • Crystallography, X-Ray / methods
  • Escherichia coli / enzymology*
  • Gene Deletion
  • Gene Expression Regulation, Enzymologic*
  • Metabolomics / methods
  • Metals / chemistry
  • Models, Chemical
  • Mutagenesis
  • Oligonucleotide Array Sequence Analysis
  • Polymers / chemistry
  • Polyribonucleotide Nucleotidyltransferase / metabolism*
  • Protein Binding

Substances

  • Metals
  • Polymers
  • Citric Acid
  • Polyribonucleotide Nucleotidyltransferase