title: Three amino acids in Escherichia coli CspE surface-exposed aromatic patch are critical for nucleic acid melting activity leading to transcription antitermination and cold acclimation of cells.
abstract: Cold-shock proteins of the CspA family of Escherichia coli help the cells to acclimate to low temperature conditions through an unknown mechanism. In vitro, these proteins bind to single-stranded nucleic acids and destabilize nucleic acid secondary structures. An unusual surface-exposed patch of 6 evolutionarily conserved aromatic amino acids is thought to be involved in RNA binding by the cold-shock proteins. Here we investigated the functional role of the aromatic patch in E. coli CspE by substituting individual aromatic residues with positively charged Arg residues. These substitutions do not affect the RNA binding activity of the CspE mutants. We show that substitutions of three centrally located aromatic patch amino acid residues, Phe(17), Phe(30), and His(32), abolish the ability of the mutant CspE to acclimatize cells to cold, antiterminate transcription and melt nucleic acids but have no effect on RNA binding. On the other hand, peripherally located Trp(10), Phe(19), and Phe(33) can be substituted with Arg without loss of any of the in vivo and in vitro CspE functions tested. The results thus indicate that these aromatic patch residues have clearly distinct functional roles and further extend the correlation between the essential function of CspA homologues in cold acclimation and their ability to antiterminate transcription.
mesh_terms: D000222:Adaptation, Physiological / Q000502:physiology*; D000596:Amino Acids / Q000502:physiology*; D001483:Base Sequence; D003080:Cold Temperature; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009696:Nucleic Acids / Q000378:metabolism*; D011487:Protein Conformation; D012329:RNA, Bacterial; D014158:Transcription, Genetic / Q000502:physiology*
chemical_list: D000596:Amino Acids; C088097:CspE protein, E coli; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D009696:Nucleic Acids; D012329:RNA, Bacterial