Transient protein interactions studied by NMR spectroscopy: the case of cytochrome C and adrenodoxin.
The interaction between yeast iso-1-cytochrome c (C102T) and two forms of bovine adrenodoxin, the wild type and a truncated form comprising residues 4-108, has been investigated using a combination of one- and two-dimensional heteronuclear NMR spectroscopy. Chemical shift perturbations and line broadening of amide resonances in the [(15)N,(1)H]HSQC spectrum for both (15)N-labeled cytochrome c and adrenodoxin in the presence of the unlabeled partner protein indicate the formation of a transient complex, with a K(a) of (4 +/- 1) x 10(4) M(-)(1) and a lifetime of <3 ms. The perturbed residues map over a large surface area for both proteins. For cytochrome c, the dominating effects are located around the exposed heme edge but with other areas also affected upon formation of the complex. In the case of adrenodoxin, effects are seen in both the recognition and core domains, with the largest perturbations in the recognition domain. These results indicate that the complex has a dynamic nature, with delocalized binding of cytochrome c on adrenodoxin. A comparison with other transient complexes of redox proteins places this complex between well-defined complexes such as the cytochrome c-cytochrome c peroxidase complex and entirely dynamic complexes such as the cytochrome b(5)-myoglobin complex.
Worrall
Jonathan A R
JA
Gorlaeus Laboratories, Leiden Institute of Chemistry, Leiden University, P.O. Box 9502, 2300 RA Leiden, The Netherlands.
Reinle
Wolfgang
W
Bernhardt
Rita
R
Ubbink
Marcellus
M
D000325:Adrenodoxin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000577:Amides / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006418:Heme / Q000737:chemistry; D057927:Hydrophobic and Hydrophilic Interactions; D007527:Isoenzymes / Q000378:metabolism; D008958:Models, Molecular; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D017186:Titrimetry / Q000379:methods
D000577:Amides; D003574:Cytochrome c Group; D007527:Isoenzymes; D009587:Nitrogen Isotopes; D000325:Adrenodoxin; D006418:Heme