The compact and expanded denatured conformations of apomyoglobin in the methanol-water solvent.
We have performed a detailed study of methanol-induced conformational transitions of horse heart apomyoglobin (apoMb) to investigate the existence of the compact and expanded denatured states. A combination of far- and near-ultraviolet circular dichroism, NMR spectroscopy, and small-angle X-ray scattering (SAXS) was used, allowing a phase diagram to be constructed as a function of pH and the methanol concentration. The phase diagram contains four conformational states, the native (N), acid-denatured (U(A)), compact denatured (I(M)), and expanded helical denatured (H) states, and indicates that the compact denatured state (I(M)) is stable under relatively mild denaturing conditions, whereas the expanded denatured states (U(A) and H) are realized under extreme conditions of pH (strong electric repulsion) or alcohol concentration (weak hydrophobic interaction). The results of this study, together with many previous studies in the literature, indicate the general existence of the compact denatured states not only in the salt-pH plane but also in the alcohol-pH plane. Furthermore, to determine the general feature of the H conformation we used several proteins including ubiquitin, ribonuclease A, alpha-lactalbumin, beta-lactoglobulin, and Streptomyces subtilisin inhibitor (SSI) in addition to apoMb. SAXS studies of these proteins in 60% methanol showed that the H states of these all proteins have expanded and nonglobular conformations. The qualitative agreement of the experimental data with computer-simulated Kratky profiles also supports this structural feature of the H state.
Kamatari
Y O
YO
The Graduate School of Science and Technology, Kobe University, Japan.
Ohji
S
S
Konno
T
T
Seki
Y
Y
Soda
K
K
Kataoka
M
M
Akasaka
K
K
D001059:Apoproteins / Q000737:chemistry*; D002942:Circular Dichroism; D003198:Computer Simulation; D004305:Dose-Response Relationship, Drug; D006863:Hydrogen-Ion Concentration; D007768:Lactalbumin / Q000737:chemistry; D007782:Lactoglobulins / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D000432:Methanol / Q000737:chemistry; D009211:Myoglobin / Q000737:chemistry*; D011487:Protein Conformation; D011489:Protein Denaturation; D012259:Ribonuclease, Pancreatic / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D014452:Ubiquitins / Q000737:chemistry; D014867:Water / Q000737:chemistry
D001059:Apoproteins; D007782:Lactoglobulins; D009211:Myoglobin; D012997:Solvents; D014452:Ubiquitins; C010857:apomyoglobin; D014867:Water; D007768:Lactalbumin; D012259:Ribonuclease, Pancreatic; D000432:Methanol