E. coli NusA protein binds in vitro to an RNA sequence immediately upstream of the boxA signal of bacteriophage lambda.

摘要: The NusA protein of Escherichia coli is a factor which mediates termination transcription. In this paper, we demonstrate that the can bind in vitro to specific site on mRNA bacteriophage lambda. Several RNAs were synthesized by transcription truncated lambda DNA templates, and activity binding these was examined Millipore filter-binding assay. containing sequence immediately upstream boxA trapped filter association with protein, but those lacking not. Anti-NusA antibody inhibits binding. To determine precisely, developed new method have named 'reverse-transcriptase mapping'. RNA transcribed from pL promoter incubated 32P-labelled primer NusA, primer-extension reaction started adding reverse transcriptase. way, extension blocked at position G (5'CGCUCUUA 3'), indicating NusA-protection includes 5'-end C. purified temperature-sensitive nusA mutant defective showed reduced thermolabile RNA-binding activity, suggesting related physiological function NusA.