Two-dimensional 1H NMR studies of cytochrome c: hydrogen exchange in the N-terminal helix
The hydrogen exchange behavior of the N-terminal helical segment in horse heart cytochrome c was studied in both the reduced and the oxidized forms by use of two-dimensional nuclear magnetic resonance methods. The amide protons of the first six residues are not H bonded and exchange rapidly with solvent protons. The most N-terminal H-bonded groups--the amide NH of Lys-7 to Phe-10--exhibit a sharp gradient in exchange rate indicative of dynamic fraying behavior, consistent with statistical-mechanical principles. This occurs identically in both reduced and oxidized cytochrome c. In the oxidized form, residues 11-14, which form the last helical turn, all exchange with a similar rate, about one million times slower than the rate characteristic of freely exposed peptide NH, even though some are on the aqueous face of the helix and others are fully buried. These and similar observations in several other proteins appear to document local cooperative unfolding reactions as determinants of protein H exchange reactions. The N-terminal segment of cytochrome c is insensitive to the heme redox state, as in the crystallographic model, except for residues closest to the heme (Cys-14 and Ala-15), which exchange about 15-fold more slowly in the reduced form. The cytochrome c H exchange results can be further considered in terms of the conformation of the native and the transiently unfolded forms and their free energy relationships in both the reduced and the oxidized states.
- Research Organization:
- Univ. of Pennsylvania, Philadelphia
- OSTI ID:
- 5698391
- Journal Information:
- Biochemistry; (United States), Vol. 5
- Country of Publication:
- United States
- Language:
- English
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ORGANIC
PHYSICAL AND ANALYTICAL CHEMISTRY
59 BASIC BIOLOGICAL SCIENCES
CYTOCHROMES
NUCLEAR MAGNETIC RESONANCE
BIOCHEMICAL REACTION KINETICS
HORSES
HYDROGEN
MYOCARDIUM
REDOX REACTIONS
THERMODYNAMICS
ANIMALS
BODY
CARDIOVASCULAR SYSTEM
CHEMICAL REACTIONS
ELEMENTS
HEART
KINETICS
MAGNETIC RESONANCE
MAMMALS
MUSCLES
NONMETALS
ORGANIC COMPOUNDS
ORGANS
PIGMENTS
PROTEINS
REACTION KINETICS
RESONANCE
VERTEBRATES
400101* - Activation
Nuclear Reaction
Radiometric & Radiochemical Procedures
550201 - Biochemistry- Tracer Techniques