EFFECT OF PEGYLATION ON THE STRUCTURE AND FUNCTION OF HORSE CYTOCHROME-C

The preparation and spectrophotometric characterization of (both Fe2and Fe3+ forms) poly(ethylene glycol) (PEG; av FW 5000)-modified horse cytochrome c (cyt c(PEG)n) with different degrees of modification (n( av) = 6, 19) by UV-vis spectroscopy, circular dichroism spectroscopy, resonance Raman spectroscopy, and cyclic voltammetry are described. Ex tensive modification (n(av) = 19) of cyt c causes gross structural def ormation of the heme as evidenced by major spectral changes in the UV- vis and circular dichroism spectral signatures of both the Fe2+ and Fe 3+ forms. Modification of cyt c by six PEG residues, however, produces a protein in which the heme active site is structurally and functiona lly intact (UV-vis, circular dichroism, and resonance Raman) and which exhibits at least quasireversible direct electron transfer (E-degrees ' = 338 +/- 5 mV vs SHE; (2.1 +/- 0.6) x 10(-3) cm/s) at bis(4-pyridyl ) disulfide-modified Au electrodes.