In vitro processing activity of Bacillus subtilis polynucleotide phosphorylase

Sutapa Mitra, Kim Hue, David H. Bechhofer

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30 Scopus citations

Abstract

A phosphate-dependent exonuclease activity was identified in purified protein fractions from Bacillus subtilis that were selected for binding to poly(I)-poly(C) agarose. Based on the characteristics of the degradation products and the absence of this activity in a pnpA strain, which contains a transposon insertion in the B. subtilis PNPase gene (Luttinger et al., 1996 - accompanying paper), this exonuclease activity was shown to be due to polynucleotide phosphorylase (PNPase). Processive 3'-to-5' exonucleolytic degradation of an SP82 phage RNA substrate was stalled at a particular site. Structure probing of the RNA showed that the stall site was downstream of a particular stem-loop structure. A similar stall site was observed for an RNA that comprised the intergenic region between the B. subtilis rpsO and pnpA genes. The ability to initiate degradation of a substrate that had a stem structure at its 3' end differed for the B. subtilis and Escherichia coli PNPase enzymes.

Original languageEnglish
Pages (from-to)329-342
Number of pages14
JournalMolecular Microbiology
Volume19
Issue number2
DOIs
StatePublished - 1996
Externally publishedYes

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