Conformational stability of apoflavodoxin.
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.
Genzor
C G
CG
Departamento de Bioquimica y Biología Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, Spain.
Beldarraín
A
A
Gómez-Moreno
C
C
López-Lacomba
J L
JL
Cortijo
M
M
Sancho
J
J
D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D001059:Apoproteins / Q000737:chemistry* / Q000378:metabolism; D002152:Calorimetry, Differential Scanning; D005418:Flavodoxin / Q000737:chemistry* / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D011487:Protein Conformation; D011489:Protein Denaturation; D014508:Urea / Q000737:chemistry
D000817:Anilino Naphthalenesulfonates; D001059:Apoproteins; D005418:Flavodoxin; D005456:Fluorescent Dyes; C026932:apoflavodoxin; C027132:1-anilino-8-naphthalenesulfonate; D014508:Urea