title: RNase E activity is conferred by a single polypeptide: overexpression, purification, and properties of the ams/rne/hmp1 gene product.
abstract: Ribonuclease E, an enzyme that processes pre-5S rRNA from its precursor, is now believed to be the major endoribonuclease participating in mRNA turnover in Escherichia coli. The product of the ams/rne/hmp1 gene, which is required for RNase E activity, was overexpressed, purified to near homogeneity by electroelution from an SDS/polyacrylamide gel, and renatured. The purified polypeptide possesses nucleolytic activity in vitro with a specificity identical to that observed for crude RNase E preparations. In addition, both UV crosslinking and RNA-protein blotting unambiguously showed that the Ams/Rne/Hmp1 polypeptide has a high affinity for RNA. Our results demonstrate that RNase E activity is directly attributable to, and is an inherent property of, an RNA-binding protein, the ams/rne/hmp1 gene product.
authors.affiliation: Department of Biochemistry, University of Western Ontario, London, Canada.
authors.forename: R S
authors.initials: RS
authors.lastname: Cormack
authors.forename: J L
authors.initials: JL
authors.lastname: Genereaux
authors.forename: G A
authors.initials: GA
authors.lastname: Mackie
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004722:Endoribonucleases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D017931:DNA Primers; D011994:Recombinant Proteins; D004722:Endoribonucleases; C022537:ribonuclease E