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The effects of amino acids substitution on electron transfer rates in mixed-metal hemoglobin hybrids
REDOX BIOCHEMISTRY
213
027
BINDING OF PORPHYRIN cmoc~m4~ C’S AND cm~m c PER~xIDASE. A.M. English, J.A. Kornblatt, Concordia University, Montreal, Quebec, Canada, H3G lM8. Iron-free, fluorescent porphyrin cytcchrome.c's from yeast (Saccharanyces cerevisiae and Candida krusei), horse and tuna have beenprepared and analyzed by gel and ion-exchange chromatography. Their interaction with ferricytochrcme c peroxidase (CCP) from baker's yeast has been monitored by fluorescence quenching and polarization. Fluorescence quenching of porphyrin c by CCP occurs on carrplexation of the two proteins, and data will be presented for different phosphate concentrations (PH 7). The extent of polarization of the porphyrin c fluorescence depends on the rotational freedom of porphyrin c when corrplexed to CCP. Again polarization data will be presented vs. phosphate concentration.
REGULATION OF INTERPROTEIN ELECTRON TRANSFER WITHIN THE Zn- CYTOCHROME C PEROXIDASE/CYTOCHROME C COMPLEX BY A CONFORMATIONAL GATE, J.M. Nocek, E.D.A. Stemp, A.G. Mauk," E. Margoliash, and B.M. Hoffman, Northwestern University, Evanston, IL 60208, USA, and *University of British Columbia, Vancouver, B.C. V6T lW5 Canada. 1ZnP + Fe3+P electron transfer within the electrostatic complex between Zn-substituted cytochrome E peroxidase and yeast (T-102) cytochrome E is abruptly abolished by cooling through the temperature range of 230-250K. This gating is modelled with a two-state mechanism [l] where for T 2 250K the complex adopts a conformation in which electron transfer is rate-limiting. Between 230 and 250K, the protein converts to a state that does not undergo electron transfer. [l] Hoffman and Ratner, J. Am. Chem. Sot., 109, 6237 (1987).
Go28
THE EFFECTS OF AMINO ACID SUBSTITUTION ON ELECTRON TRANSFER RATES IN MIXED-METAL HEMOGLOBIN HYBRIDS. D. J. Ginerich, B. M. Hoffman, Northwestern University, Evanston, IL 60208, USA. Long-range electron transfer studies in mixed-metal hybrid hemoglobins have been expanded to include the effects of the protein environment between the donor and acceptor hemes by preparing hybrids using the BFe subunits of naturally occurring hemoglobin variants and the metal-substituted u chains of normal A0 hemoglobin. To date, these variants include Hb Kempsey (B99 ASP + ASN) and Hb Yakima (/?99ASP -+ HIS). Both the photochemical forward (triplet to Fe(II1) heme) and thermal back (Fe(II1) heme to the Zn or Mg n-cation radical) reactions can be monitored to assess the role of individual amino acid residues.
GO29
213
027
BINDING OF PORPHYRIN cmoc~m4~ C’S AND cm~m c PER~xIDASE. A.M. English, J.A. Kornblatt, Concordia University, Montreal, Quebec, Canada, H3G lM8. Iron-free, fluorescent porphyrin cytcchrome.c's from yeast (Saccharanyces cerevisiae and Candida krusei), horse and tuna have beenprepared and analyzed by gel and ion-exchange chromatography. Their interaction with ferricytochrcme c peroxidase (CCP) from baker's yeast has been monitored by fluorescence quenching and polarization. Fluorescence quenching of porphyrin c by CCP occurs on carrplexation of the two proteins, and data will be presented for different phosphate concentrations (PH 7). The extent of polarization of the porphyrin c fluorescence depends on the rotational freedom of porphyrin c when corrplexed to CCP. Again polarization data will be presented vs. phosphate concentration.
REGULATION OF INTERPROTEIN ELECTRON TRANSFER WITHIN THE Zn- CYTOCHROME C PEROXIDASE/CYTOCHROME C COMPLEX BY A CONFORMATIONAL GATE, J.M. Nocek, E.D.A. Stemp, A.G. Mauk," E. Margoliash, and B.M. Hoffman, Northwestern University, Evanston, IL 60208, USA, and *University of British Columbia, Vancouver, B.C. V6T lW5 Canada. 1ZnP + Fe3+P electron transfer within the electrostatic complex between Zn-substituted cytochrome E peroxidase and yeast (T-102) cytochrome E is abruptly abolished by cooling through the temperature range of 230-250K. This gating is modelled with a two-state mechanism [l] where for T 2 250K the complex adopts a conformation in which electron transfer is rate-limiting. Between 230 and 250K, the protein converts to a state that does not undergo electron transfer. [l] Hoffman and Ratner, J. Am. Chem. Sot., 109, 6237 (1987).
Go28
THE EFFECTS OF AMINO ACID SUBSTITUTION ON ELECTRON TRANSFER RATES IN MIXED-METAL HEMOGLOBIN HYBRIDS. D. J. Ginerich, B. M. Hoffman, Northwestern University, Evanston, IL 60208, USA. Long-range electron transfer studies in mixed-metal hybrid hemoglobins have been expanded to include the effects of the protein environment between the donor and acceptor hemes by preparing hybrids using the BFe subunits of naturally occurring hemoglobin variants and the metal-substituted u chains of normal A0 hemoglobin. To date, these variants include Hb Kempsey (B99 ASP + ASN) and Hb Yakima (/?99ASP -+ HIS). Both the photochemical forward (triplet to Fe(II1) heme) and thermal back (Fe(II1) heme to the Zn or Mg n-cation radical) reactions can be monitored to assess the role of individual amino acid residues.
GO29