title: Conformational states of beta-lactamase: molten-globule states at acidic and alkaline pH with high salt.
abstract: We present evidence that beta-lactamase is close to fully unfolded (i.e., random coil conformation) at low ionic strength at the extremes of pH and that the presence of salt causes a cooperative transition to a conformation with the properties of a molten globule, namely, a compact state with native-like secondary structure but disordered side chains (tertiary structure). The conformation of beta-lactamase I from Bacillus cereus was examined over the pH 1.5-12.5 region by circular dichroism (CD), tryptophan fluorescence, dynamic light scattering, and 1-anilino-8-naphthalenesulfonate (ANS) binding. Under conditions of low ionic strength (I = 0.05) beta-lactamase was unfolded below pH 2.5 and above pH 11.5, on the basis of the far-UV and near-UV CD and tryptophan fluorescence. However, at high ionic strength and low pH an intermediate conformation (state A) was observed, with a secondary structure content similar to that of the native protein but a largely disordered tertiary structure. The transition from the unfolded state (U) to state A induced by KCl was cooperative and had a midpoint at 0.12 M KCl (I = 0.17 M) at pH 1.6. A similar conformation (state B) was observed at high pH and high ionic strength. The transition from the alkaline U state to state B induced by KCl at pH 12.2 was cooperative and had a midpoint at 0.6 M KCl (I = 0.65 M). Light scattering measurements showed that state B was compact although somewhat expanded compared to the N state. The compactness of state A could not be determined due to its strong propensity to aggregate.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Chemistry, University of California, Santa Cruz 95064.
authors.forename: Y
authors.initials: Y
authors.lastname: Goto
authors.forename: A L
authors.initials: AL
authors.lastname: Fink
mesh_terms: D001409:Bacillus cereus / Q000201:enzymology; D002942:Circular Dichroism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009994:Osmolar Concentration; D010405:Penicillinase / Q000378:metabolism*; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
chemical_list: D010405:Penicillinase