microbiology, Vol 142, 359-365, Copyright © 1996 by Society for General Microbiology

ARTICLES

The defective phosphoribosyl diphosphate synthase in a temperature- sensitive prs-2 mutant of Escherichia coli is compensated by increased enzyme synthesis

DA Post, RL Switzer and B Hove-Jensen
Department of Biochemistry, University of Illinois, Urbana 61801, USA.

An Escherichia coli strain which is temperature-sensitive for growth due to a mutation (prs-2) causing a defective phosphoribosyl diphosphate (PRPP) synthase has been characterized. The temperature- sensitive mutation was mapped to a 276 bp HindIII-BssHII DNA fragment located within the open reading frame specifying the PRPP synthase polypeptide. Cloning and sequencing of the mutant allele revealed two mutations. One, a G --> A transition, located in the ninth codon, was responsible for the temperature-conditional phenotype and resulted in a serine residue at this position. The wild-type codon at this position specified a glycine residue that is conserved among PRPP synthases across a broad phylogenetic range. Cells harbouring the glycine-to- serine alteration specified by a plasmid contained approximately 50% of the PRPP synthase activity of cells harbouring a plasmid-borne wild- type allele, both grown at 25 degrees C. The mutant enzyme had nearly normal heat stability, as long as it was synthesized at 25 degrees C. In contrast, there was hardly any PRPP synthase activity or anti-PRPP synthase antibody cross-reactive material present in cells harbouring the glycine to serine alteration following temperature shift to 42 degrees C. The other mutation was a C --> T transition located 39 bp upstream of the G --> A mutation, i.e. outside the coding sequence and close to the Shine-Dalgarno sequence. Cells harbouring only the C --> T mutation in a plasmid contained approximately three times as much PRPP synthase activity as a strain harbouring a plasmid-borne wild-type prs allele. In cells harbouring both mutations, the C --> T mutation appeared to compensate for the G --> A mutation by increasing the amount of a partially defective enzyme at the permissive temperature.

This article has been cited by other articles:

				B. Hove-Jensen Heterooligomeric Phosphoribosyl Diphosphate Synthase of Saccharomyces cerevisiae: COMBINATORIAL EXPRESSION OF THE FIVE PRS GENES IN ESCHERICHIA COLI J. Biol. Chem., September 24, 2004; 279(39): 40345 - 40350. [Abstract] [Full Text] [PDF]

				B. Hove-Jensen, T. J. Rosenkrantz, A. Haldimann, and B. L. Wanner Escherichia coli phnN, Encoding Ribose 1,5-Bisphosphokinase Activity (Phosphoribosyl Diphosphate Forming): Dual Role in Phosphonate Degradation and NAD Biosynthesis Pathways J. Bacteriol., May 1, 2003; 185(9): 2793 - 2801. [Abstract] [Full Text] [PDF]

				A. W. Karzai and R. T. Sauer Protein factors associated with the SsrA{middle dot}SmpB tagging and ribosome rescue complex PNAS, February 22, 2001; (2001) 51628298. [Abstract] [Full Text]

				B. N. Krath and B. Hove-Jensen Organellar and Cytosolic Localization of Four Phosphoribosyl Diphosphate Synthase Isozymes in Spinach Plant Physiology, February 1, 1999; 119(2): 497 - 506. [Abstract] [Full Text]

				B. N. Krath and B. Hove-Jensen Class II Recombinant Phosphoribosyl Diphosphate Synthase from Spinach. PHOSPHATE INDEPENDENCE AND DIPHOSPHORYL DONOR SPECIFICITY J. Biol. Chem., May 18, 2001; 276(21): 17851 - 17856. [Abstract] [Full Text] [PDF]

				A. W. Karzai and R. T. Sauer Protein factors associated with the SsrA{middle dot}SmpB tagging and ribosome rescue complex PNAS, March 13, 2001; 98(6): 3040 - 3044. [Abstract] [Full Text] [PDF]