title: Folding kinetics of phage 434 Cro protein.
abstract: Folding kinetics for phage 434 Cro protein are examined and compared with those reported for lambda(6-85), the N-terminal domain of the repressor of phage lambda. The two proteins have similar all-helical structures consisting of five helices but different stabilities. In contrast to lambda(6-85), sharp and distinct aromatic (1)H NMR signals without exchange broadening characterize the native and urea-denatured 434 Cro forms at equilibrium at 20 degrees C, indicating slow interconversion on the NMR time scale. Stopped-flow fluorescence data using the single 434 Cro tryptophan indicate strongly urea-dependent refolding rates and smaller urea dependencies of the unfolding rates, suggesting a native-like transition state ensemble. Refolding rates are slower and unfolding rates considerably faster at pH 4 than at pH 6. This accounts for the lower stability of 434 Cro at pH 4 and suggests the existence of pH-dependent, possibly salt bridge interactions that are more stabilizing at pH 6. At <2 M urea, decreased folding amplitudes and nonlinear urea dependencies that are apparent at pH 6 indicate deviation from two-state behavior and suggest the formation of an early folding intermediate. The folding behavior of 434 Cro and why it folds 2 orders of magnitude slower than lambda(6-85) are rationalized in terms of the lower intrinsic helix stabilities and putative charge interactions in 434 Cro.
authors.affiliation: Instituto de Estructura de la Materia, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006 Madrid, Spain.
authors.forename: D V
authors.initials: DV
authors.lastname: Laurents
authors.forename: S
authors.initials: S
authors.lastname: Corrales
authors.forename: M
authors.initials: M
authors.lastname: Elías-Arnanz
authors.forename: P
authors.initials: P
authors.lastname: Sevilla
authors.forename: M
authors.initials: M
authors.lastname: Rico
authors.forename: S
authors.initials: S
authors.lastname: Padmanabhan
mesh_terms: D000595:Amino Acid Sequence; D010582:Bacteriophage lambda / Q000737:chemistry; D002942:Circular Dichroism; D003090:Coliphages / Q000737:chemistry*; D004268:DNA-Binding Proteins; D007700:Kinetics; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry; D011489:Protein Denaturation; D017510:Protein Folding; D012097:Repressor Proteins / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D013696:Temperature; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins
chemical_list: C063367:434-repressor protein, Bacteriophage 434; D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins