Solvent-induced ligand dissociation and conformational states of Cellular Retinol-Binding Protein type I.
Cellular Retinol-Binding Protein type I (CRBP) exhibits very high affinity for its ligand, bound within a buried cavity completely shielded from the outside medium. Three-dimensional structure and backbone dynamics in aqueous solution at neutral pH, either in the absence or in the presence of retinol, fail to represent the protein in a state capable of ligand uptake and release. The question was asked whether changes in the composition of the outside medium might facilitate ligand dissociation. Acidic aqueous solutions and water-alcohol mixtures were selected, among the best described denaturing solvents, to investigate their effects on the stability of the carrier-ligand complex and the conformational state of the protein upon ligand release. Circular dichroism (CD) and fluorescence spectroscopy were used to probe protein secondary and tertiary structure, compactness and retinol dissociation. While in purely aqueous media retinol dissociation parallels the acid-induced denaturation of the carrier, in water-alcohol mixtures it occurs in a range of co-solvent content lower than that required for protein denaturation. In light of these results, it is suggested that local solvent properties in vivo might modulate protein conformation and flexibility and thus play a fundamental role in the control of retinol exchange between carrier and membrane-bound donors and acceptors.
Torta
Federico
F
Department of Biochemistry and Molecular Biology, University of Parma, Parco Area delle Scienze 23/A, I-43100 Parma, Italy.
Dyuysekina
Alexandra E
AE
Cavazzini
Davide
D
Fantuzzi
Andrea
A
Bychkova
Valentina E
VE
Rossi
Gian Luigi
GL
D000143:Acids / Q000737:chemistry; D000438:Alcohols / Q000737:chemistry; D000818:Animals; D001059:Apoproteins / Q000737:chemistry / Q000378:metabolism; D002942:Circular Dichroism; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008024:Ligands; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects*; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats; D012177:Retinol-Binding Proteins / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D054840:Retinol-Binding Proteins, Cellular; D012997:Solvents / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D014508:Urea / Q000494:pharmacology; D014801:Vitamin A / Q000737:chemistry* / Q000378:metabolism / Q000493:pharmacokinetics; D014867:Water / Q000737:chemistry
D000143:Acids; D000438:Alcohols; D001059:Apoproteins; D008024:Ligands; D012177:Retinol-Binding Proteins; D054840:Retinol-Binding Proteins, Cellular; D012997:Solvents; D014867:Water; D014801:Vitamin A; D014508:Urea