Characterization of the stable, acid‐induced, molten globule‐like state of staphylococcal nuclease

Titration of a salt‐free solution of native staphylococcal nuclease by HC1 leads to an unfolding transition in the vicinity of pH 4, as determined by near‐ and far‐UV circular dichroism. At pH 2‐3, the protein is substantially unfolded. The addition of further HC1 results in a second transition, this one to a more structured species (the A state) with the properties of an expanded molten globule, namely substantial secondary structure, little or no tertiary structure, relatively compact size as determined by hydrodynamic radius, and the ability to bind the hydrophobic dye 1‐anilino‐8‐naphthalene sulfonic acid. The addition of anions, in the form of neutral salts, to the acid‐unfolded state at pH 2 also causes a transition leading to the A state. Fourier transform infrared analysis of the amide I band was used to compare the amount and type of secondary structure in the native and A states. A significant decrease in α‐helix structure, with a corresponding increase in β or extended structure, was observed in the A state, compared to the native state. A model to account for such compact denatured states is proposed.

[1]  D. Engelman,et al.  Truncated staphylococcal nuclease is compact but disordered. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[2]  K. Kuwajima,et al.  Folding of staphylococcal nuclease A studied by equilibrium and kinetic circular dichroism spectra. , 1991, Biochemistry.

[3]  R. L. Baldwin,et al.  Early folding intermediate of ribonuclease A. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[4]  A. Fersht,et al.  Detection and characterization of a folding intermediate in barnase by NMR , 1990, Nature.

[5]  A. Fink,et al.  Mechanism of acid-induced folding of proteins. , 1990, Biochemistry.

[6]  O. Ptitsyn,et al.  Evidence for a molten globule state as a general intermediate in protein folding , 1990, FEBS letters.

[7]  P. S. Kim,et al.  Intermediates in the folding reactions of small proteins. , 1990, Annual review of biochemistry.

[8]  A. Fink,et al.  Acid-induced folding of proteins. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[9]  D. Shortle,et al.  Effects of denaturants at low concentrations on the reversible denaturation of staphylococcal nuclease. , 1989, Archives of biochemistry and biophysics.

[10]  D. Shortle,et al.  Residual structure in large fragments of staphylococcal nuclease: effects of amino acid substitutions. , 1989, Biochemistry.

[11]  A. Fink,et al.  Conformational states of beta-lactamase: molten-globule states at acidic and alkaline pH with high salt. , 1989, Biochemistry.

[12]  K. Kuwajima,et al.  The molten globule state as a clue for understanding the folding and cooperativity of globular‐protein structure , 1989, Proteins.

[13]  S. Walter Englander,et al.  Structural characterization of folding intermediates in cytochrome c by H-exchange labelling and proton NMR , 1988, Nature.

[14]  O. Ptitsyn,et al.  Sequential mechanism of refolding of carbonic anhydrase B , 1987, FEBS letters.

[15]  O. Ptitsyn,et al.  An early intermediate of refolding α‐lactalbumin forms within 20 ms , 1987 .

[16]  S Sugai,et al.  Rapid formation of secondary structure framework in protein folding studied by stopped‐flow circular dichroism , 1987, FEBS letters.

[17]  O. Ptitsyn.  Protein folding: Hypotheses and experiments , 1987 .

[18]  R. Jakobsen,et al.  Effects of the Environment on the Structure of Adsorbed Proteins: Fourier Transform Infrared Spectroscopic Studies , 1987 .

[19]  K. Kuwajima,et al.  Influence of Ca2+ binding on the structure and stability of bovine alpha-lactalbumin studied by circular dichroism and nuclear magnetic resonance spectra. , 2009, International journal of peptide and protein research.

[20]  K. Kuwajima,et al.  Evidence for identity between the equilibrium unfolding intermediate and a transient folding intermediate: a comparative study of the folding reactions of alpha-lactalbumin and lysozyme. , 1986, Biochemistry.

[21]  H. Susi,et al.  Examination of the secondary structure of proteins by deconvolved FTIR spectra , 1986, Biopolymers.

[22]  J. Bandekar,et al.  Vibrational spectroscopy and conformation of peptides, polypeptides, and proteins. , 1986, Advances in protein chemistry.

[23]  K. Kuwajima,et al.  Comparison of the transient folding intermediates in lysozyme and alpha-lactalbumin. , 1985, Biochemistry.

[24]  R. J. Corbett,et al.  Use of high-speed size-exclusion chromatography for the study of protein folding and stability. , 1984, Biochemistry.

[25]  A. Wada,et al.  ‘Molten‐globule state’: a compact form of globular proteins with mobile side‐chains , 1983, FEBS letters.

[26]  D. Worah,et al.  Binding of recrystallized and chromatographically purified 8-anilino-1-naphthalenesulfonate to Escherichia coli lac repressor. , 1978, Biochemistry.