title: Identification and thermodynamic characterization of molten globule states of periplasmic binding proteins.
abstract: Molten globule-like intermediates have been shown to occur during protein folding and are thought to be involved in protein translocation and membrane insertion. However, the determinants of molten globule stability and the extent of specific packing in molten globules is currently unclear. Using far- and near-UV CD and intrinsic and ANS fluorescence, we show that four periplasmic binding proteins (LBP, LIVBP, MBP, and RBP) form molten globules at acidic pH values ranging from 3.0 to 3.4. Only two of these (LBP and LIVBP) have similar sequences, but all four proteins adopt similar three-dimensional structures. We found that each of the four molten globules binds to its corresponding ligand without conversion to the native state. Ligand binding affinity measured by isothermal titration calorimetry for the molten globule state of LIVBP was found to be comparable to that of the corresponding native state, whereas for LBP, MBP, and RBP, the molten globules bound ligand with approximately 5-30-fold lower affinity than the corresponding native states. All four molten globule states exhibited cooperative thermal unfolding assayed by DSC. Estimated values of DeltaCp of unfolding show that these molten globule states contain 28-67% of buried surface area relative to the native states. The data suggest that molten globules of these periplasmic binding proteins retain a considerable degree of long range order. The ability of these sequentially unrelated proteins to form highly ordered molten globules may be related to their large size as well as an intrinsic property of periplasmic binding protein folds.
authors: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India.
authors: Ravindra Singh
authors: RS
authors: Prajapati
authors: S
authors: S
authors: Indu
authors: Raghavan
authors: R
authors: Varadarajan
mesh_terms: D002152:Calorimetry, Differential Scanning; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008024:Ligands; D033902:Periplasmic Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics
chemical_list: D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D008024:Ligands; C488923:LivJ protein, E coli; D033902:Periplasmic Binding Proteins