Abstract
Bacterial small RNA molecules (sRNAs) are increasingly recognized as central regulators of bacterial stress responses and pathogenesis. In many cases, RNA-binding proteins are critical for the stability and function of sRNAs. Previous studies have adopted strategies to genetically tag an sRNA of interest, allowing isolation of RNA-protein complexes from cells. Here we present a sequence-specific affinity purification protocol that requires no prior genetic manipulation of bacterial cells, allowing isolation of RNA-binding proteins bound to native RNA molecules.
Keywords:
Affinity chromatography; Bacterial small RNAs; Hfq; In vivo crosslinking; RNA-binding proteins; Western blot.
Publication types
-
Research Support, N.I.H., Extramural
-
Research Support, Non-U.S. Gov't
MeSH terms
-
Bacterial Proteins / genetics
-
Bacterial Proteins / isolation & purification
-
Bacterial Proteins / metabolism*
-
Binding Sites
-
Chromatography, Affinity / methods*
-
Cross-Linking Reagents / metabolism
-
Protein Binding
-
Pseudomonas aeruginosa / genetics
-
Pseudomonas aeruginosa / growth & development
-
Pseudomonas aeruginosa / metabolism*
-
RNA, Bacterial / genetics
-
RNA, Bacterial / metabolism*
-
RNA, Small Untranslated / genetics
-
RNA, Small Untranslated / metabolism*
-
RNA-Binding Proteins / genetics
-
RNA-Binding Proteins / isolation & purification
-
RNA-Binding Proteins / metabolism*
-
Ultraviolet Rays
Substances
-
Bacterial Proteins
-
Cross-Linking Reagents
-
RNA, Bacterial
-
RNA, Small Untranslated
-
RNA-Binding Proteins