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Equilibrium unfolding of dimeric and engineered monomeric forms of lambda Cro (F58W) repressor and the effect of added salts: evidence for the formation of folded monomer induced by sodium perchlorate.
Arch Biochem Biophys. 2005 Feb 01; 434(1):93-107.AB

Abstract

The equilibrium unfolding transitions of Cro repressor variants, dimeric variant Cro F58W and monomer Cro K56[DGEVK]F58W, have been studied by urea and guanidine hydrochloride to probe the folding mechanism. The unfolding transitions of a dimeric variant are well described by a two state process involving native dimer and unfolded monomer with a free energy of unfolding, DeltaG(0,un)(0), of approximately 10-11 kcal/mol. The midpoint of transition curves is dependent on total protein concentration and DeltaG(0,un)(0) is independent of protein concentration, as expected for this model. Unfolding of Cro monomer is well described by the standard two state model. The stability of both forms of protein increases in the presence of salt but decreases with the decrease in pH. Because of the suggested importance of a N2<-->2F dimerization process in DNA binding, we have also studied the effect of sodium perchlorate, containing the chaotropic perchlorate anion, on the conformational transition of Cro dimer by CD, fluorescence and NMR (in addition to urea and guanidine hydrochloride) in an attempt both to characterize the thermodynamics of the process and to identify conditions that lead to an increase in the population of the folded monomers. Data suggest that sodium perchlorate stabilizes the protein at low concentration (<1.5 M) and destabilizes the protein at higher perchlorate concentration with the formation of a "significantly folded" monomer. The tryptophan residue in the "significantly folded" monomer induced by perchlorate is more exposed to the solvent than in native dimer.

Authors+Show Affiliations

Department of Chemistry and Biochemistry, University of Mississippi, Coulter Hall University, MS 38677, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.

Language

eng

PubMed ID

15629113

Citation

Maity, Haripada, et al. "Equilibrium Unfolding of Dimeric and Engineered Monomeric Forms of Lambda Cro (F58W) Repressor and the Effect of Added Salts: Evidence for the Formation of Folded Monomer Induced By Sodium Perchlorate." Archives of Biochemistry and Biophysics, vol. 434, no. 1, 2005, pp. 93-107.
Maity H, Mossing MC, Eftink MR. Equilibrium unfolding of dimeric and engineered monomeric forms of lambda Cro (F58W) repressor and the effect of added salts: evidence for the formation of folded monomer induced by sodium perchlorate. Arch Biochem Biophys. 2005;434(1):93-107.
Maity, H., Mossing, M. C., & Eftink, M. R. (2005). Equilibrium unfolding of dimeric and engineered monomeric forms of lambda Cro (F58W) repressor and the effect of added salts: evidence for the formation of folded monomer induced by sodium perchlorate. Archives of Biochemistry and Biophysics, 434(1), 93-107.
Maity H, Mossing MC, Eftink MR. Equilibrium Unfolding of Dimeric and Engineered Monomeric Forms of Lambda Cro (F58W) Repressor and the Effect of Added Salts: Evidence for the Formation of Folded Monomer Induced By Sodium Perchlorate. Arch Biochem Biophys. 2005 Feb 1;434(1):93-107. PubMed PMID: 15629113.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Equilibrium unfolding of dimeric and engineered monomeric forms of lambda Cro (F58W) repressor and the effect of added salts: evidence for the formation of folded monomer induced by sodium perchlorate. AU - Maity,Haripada, AU - Mossing,Michael C, AU - Eftink,Maurice R, PY - 2004/09/02/received PY - 2005/1/5/pubmed PY - 2005/2/11/medline PY - 2005/1/5/entrez SP - 93 EP - 107 JF - Archives of biochemistry and biophysics JO - Arch Biochem Biophys VL - 434 IS - 1 N2 - The equilibrium unfolding transitions of Cro repressor variants, dimeric variant Cro F58W and monomer Cro K56[DGEVK]F58W, have been studied by urea and guanidine hydrochloride to probe the folding mechanism. The unfolding transitions of a dimeric variant are well described by a two state process involving native dimer and unfolded monomer with a free energy of unfolding, DeltaG(0,un)(0), of approximately 10-11 kcal/mol. The midpoint of transition curves is dependent on total protein concentration and DeltaG(0,un)(0) is independent of protein concentration, as expected for this model. Unfolding of Cro monomer is well described by the standard two state model. The stability of both forms of protein increases in the presence of salt but decreases with the decrease in pH. Because of the suggested importance of a N2<-->2F dimerization process in DNA binding, we have also studied the effect of sodium perchlorate, containing the chaotropic perchlorate anion, on the conformational transition of Cro dimer by CD, fluorescence and NMR (in addition to urea and guanidine hydrochloride) in an attempt both to characterize the thermodynamics of the process and to identify conditions that lead to an increase in the population of the folded monomers. Data suggest that sodium perchlorate stabilizes the protein at low concentration (<1.5 M) and destabilizes the protein at higher perchlorate concentration with the formation of a "significantly folded" monomer. The tryptophan residue in the "significantly folded" monomer induced by perchlorate is more exposed to the solvent than in native dimer. SN - 0003-9861 UR - https://www.unboundmedicine.com/medline/citation/15629113/Equilibrium_unfolding_of_dimeric_and_engineered_monomeric_forms_of_lambda_Cro__F58W__repressor_and_the_effect_of_added_salts:_evidence_for_the_formation_of_folded_monomer_induced_by_sodium_perchlorate_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0003-9861(04)00577-6 DB - PRIME DP - Unbound Medicine ER -