Cross-linked metalloproteins: Novel systems for the study of intraprotein electron-transfer reactions
My initial research shows, for the first time, that proteins can be cross-linked selectively via transition-metal compounds to form stable protein complexes. Incubation of horse heat cytochrome c (designated cyt) with reagents PtCl{sub 4}{sup 2{minus}} and trans-(Pt(2-Fpy){sub 2}Cl{sub 2}) under mild conditions yields stable diprotein complexes trans-(PtCl{sub 2}(cyt){sub 2}) and trans-(Pt(2-Fpy){sub 2}(cyt){sub 2}), respectively (2-Fpy is 2-fluoropyridine). The complexes are purified and characterized chromatographically. Spectroscopic and electrochemical measurements indicate that the structural and redox properties of the cytochrome c molecules remain virtually unaltered upon cross-linking. The diprotein complexes are stable indefinitely under ordinary conditions and yet they can be cleaved, and the native protein restored, in a mild reaction. Subsequent research focused on oxidoreduction reactions involving the electrostatic and the covalent complex of horse heart cytochrome c and Phaseolus vulgaris plastocyanin, and exploring the importance of the protein rearrangement for the intracomplex electron-transfer reaction. Cytochrome c and plastocyanin are cross-linked one-to-one by a carbodiimide, in the same general orientation in which they associate electrostatically. Four isomers of the covalent diprotein complex, which probably differ slightly from one another in the manner of cross-linking, are separated efficiently by cation-exchange chromatography. Electron transfer kinetics of the covalent and electrostatic diprotein complexes are studied using stopped-flow, pulse radiolysis, and flash photolysis experiments. 135 refs., 5 figs., 15 tabs.
- Research Organization:
- Ames Lab., IA (USA)
- Sponsoring Organization:
- DOE/ER
- DOE Contract Number:
- W-7405-ENG-82
- OSTI ID:
- 7118554
- Report Number(s):
- IS-T-1433; ON: DE90011814
- Country of Publication:
- United States
- Language:
- English
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59 BASIC BIOLOGICAL SCIENCES
ANIMALS
AROMATICS
CHEMICAL RADIATION EFFECTS
CHEMICAL REACTION KINETICS
CHEMICAL REACTIONS
CHEMISTRY
CHLORINE COMPOUNDS
CLEAVAGE
COMPLEXES
CROSS-LINKING
CRYSTAL STRUCTURE
CYTOCHROMES
DECOMPOSITION
ELECTRON TRANSFER
HALOGEN COMPOUNDS
HETEROCYCLIC COMPOUNDS
HORSES
IRON COMPLEXES
ISOALLOXAZINES
ISOMERS
KINETICS
MAMMALS
METALLOPROTEINS
MICROSTRUCTURE
MOLECULAR STRUCTURE
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC OXYGEN COMPOUNDS
OXIDIZERS
PHOTOCHEMICAL REACTIONS
PHOTOLYSIS
PIGMENTS
PLATINUM COMPLEXES
POLYMERIZATION
PROTEIN STRUCTURE
PROTEINS
QUINONES
RADIATION CHEMISTRY
RADIATION EFFECTS
RADIOLYSIS
REACTION KINETICS
REDOX REACTIONS
REDUCING AGENTS
SPECIFICITY
TRANSITION ELEMENT COMPLEXES
VERTEBRATES