Conformational stability of apoflavodoxin.


Abstract

Flavodoxins are alpha/beta proteins that mediate electron transfer reactions. The conformational stability of apoflavodoxin from Anaboena PCC 7119 has been studied by calorimetry and urea denaturation as a function of pH and ionic strength. At pH > 12, the protein is unfolded. Between pH 11 and pH 6, the apoprotein is folded properly as judged from near-ultraviolet (UV) circular dichroism (CD) and high-field 1H NMR spectra. In this pH interval, apoflavodoxin is a monomer and its unfolding by urea or temperature follows a simple two-state mechanism. The specific heat capacity of unfolding for this native conformation is unusually low. Near its isoelectric point (3.9), the protein is highly insoluble. At lower pH values (pH 3.5-2.0), apoflavodoxin adopts a conformation with the properties of a molten globule. Although apoflavodoxin at pH 2 unfolds cooperatively with urea in a reversible fashion and the fluorescence and far-UV CD unfolding curves coincide, the transition midpoint depends on the concentration of protein, ruling out a simple two-state process at acidic pH. Apoflavodoxin constitutes a promising system for the analysis of the stability and folding of alpha/beta proteins and for the study of the interaction between apoflavoproteins and their corresponding redox cofactors. Study holds ProTherm entries: 8639, 8640, 8641, 8642, 8643, 8644, 8645, 8646, 8647, 8648, 8649 Extra Details: calorimetry; flavodoxin; molten globule; protein folding;,protein stability; redox proteins

Submission Details

ID: WUaA7jec

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:36 p.m.

Version: 1

Publication Details
Genzor CG;Beldarraín A;Gómez-Moreno C;López-Lacomba JL;Cortijo M;Sancho J,Protein Sci. (1996) Conformational stability of apoflavodoxin. PMID:8819170
Additional Information

Number of data points 38
Proteins Flavodoxin ; Flavodoxin
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: dCp units:kJ/mol·K, buffers:Sodium phosphate: 50 mM, temp:25.0 C, pH:7.0 ; Experimental Assay: dHcal buffers:Sodium phosphate: 50 mM, temp:25.0 C, pH:7.0 ; Experimental Assay: dG ; Experimental Assay: dHcal buffers:glycine: 50 mM, pH:11.0 ; Experimental Assay: Tm buffers:glycine: 50 mM, pH:11.0 ; Experimental Assay: dCp buffers:glycine: 50 mM, pH:10.0, units:kJ/mol ; Experimental Assay: dHcal buffers:glycine: 50 mM, pH:10.0 ; Experimental Assay: Tm buffers:glycine: 50 mM, pH:10.0 ; Experimental Assay: dHvH buffers:glycine: 50 mM, pH:10.0 ; Experimental Assay: dCp buffers:phosphate: 50 mM, pH:9.0, units:kJ/mol ; Experimental Assay: dHcal buffers:phosphate: 50 mM, pH:9.0 ; Experimental Assay: Tm buffers:phosphate: 50 mM, pH:9.0 ; Experimental Assay: dHvH buffers:phosphate: 50 mM, pH:9.0 ; Experimental Assay: dCp buffers:phosphate: 50 mM, pH:8.0, units:kJ/mol ; Experimental Assay: dHcal buffers:phosphate: 50 mM, pH:8.0 ; Experimental Assay: Tm buffers:phosphate: 50 mM, pH:8.0 ; Experimental Assay: dHvH buffers:phosphate: 50 mM, pH:8.0 ; Experimental Assay: dCp buffers:phosphate: 20 mM, pH:8.0, units:kJ/mol ; Experimental Assay: dHcal buffers:phosphate: 20 mM, pH:8.0 ; Experimental Assay: Tm buffers:phosphate: 20 mM, pH:8.0 ; Experimental Assay: dHvH buffers:phosphate: 20 mM, pH:8.0 ; Experimental Assay: dCp pH:7.0, buffers:phosphate: 50 mM, units:kJ/mol ; Experimental Assay: dHcal buffers:phosphate: 50 mM, pH:7.0 ; Experimental Assay: Tm buffers:phosphate: 50 mM, pH:7.0 ; Experimental Assay: dHvH buffers:phosphate: 50 mM, pH:7.0 ; Experimental Assay: dCp pH:7.0, buffers:phosphate: 20 mM, units:kJ/mol ; Experimental Assay: dHcal buffers:phosphate: 20 mM, pH:7.0 ; Experimental Assay: Tm buffers:phosphate: 20 mM, pH:7.0 ; Experimental Assay: dHvH buffers:phosphate: 20 mM, pH:7.0 ; Experimental Assay: dCp pH:6.0, buffers:phosphate: 50 mM, units:kJ/mol ; Experimental Assay: dHcal pH:6.0, buffers:phosphate: 50 mM ; Experimental Assay: Tm pH:6.0, buffers:phosphate: 50 mM ; Experimental Assay: dHvH pH:6.0, buffers:phosphate: 50 mM ; Experimental Assay: dCp pH:6.0, buffers:sodium phosphate: 20 mM, units:kJ/mol ; Experimental Assay: dHcal pH:6.0, buffers:sodium phosphate: 20 mM ; Experimental Assay: Tm pH:6.0, buffers:sodium phosphate: 20 mM ; Experimental Assay: dHvH pH:6.0, buffers:sodium phosphate: 20 mM ; Experimental Assay: dG_H2O
Libraries Mutations for sequence KKIGLFYGTQTGKTESVAEIIRDEFGNDVVTLHDVSQAEVTDLNDYQYLIIGCPTWNIGELQSDWEGLYSELDDVDFNGKLVAYFGTGDQIGYADNFQDAIGILEEKISQRGGKTVGYWSTDGYDFNDSKALRNGKFVGLALDEDNQSDLTDDRIKSWVAQLKSEFGL
Sequence Assay Result Units