Isolation and characterization of a new temperature-sensitive polynucleotide phosphorylase mutation in Escherichia coli K-12

• Published in: Biochimie Vol. 72; no. 11; p. 835
• Main Authors: Yancey, S D, Kushner, S R
• Format: Journal Article
• Language: English
• Published: France 01-11-1990
• Subjects: Escherichia coli - enzymology; Escherichia coli - genetics; Genes, Bacterial; Genotype; Hot Temperature; Kinetics; Mutation; Plasmids; Polyribonucleotide Nucleotidyltransferase - genetics; Polyribonucleotide Nucleotidyltransferase - isolation & purification; Polyribonucleotide Nucleotidyltransferase - metabolism; Restriction Mapping; Thermodynamics
• ISSN: 0300-9084
• DOI: 10.1016/0300-9084(90)90193-K

Polynucleotide phosphorylase (PNPase) has been studied in detail since its discovery in 1955 [1]. In an attempt to determine what role, if any, it has in mRNA decay in Escherichia coli, we have isolated and characterized a temperature-sensitive mutation, pnp-200, in the pnp gene. In vitro phosphorolysis, polymerization and exchange activities of the partially purified Pnp-200 enzyme are all reduced to 30-40% of wild-type activity at 50 degrees C compared to 32 degrees C. The pnp-200 mutation alone does not affect cell growth or mRNA stability. A triple mutant strain containing pnp-200 in combination with other temperature-sensitive mutations in genes known to affect mRNA metabolism (rnb-500 and ams-1) is conditionally lethal and shows increased mRNA stability after shift to the non-permissive temperature.