KINETICS OF THE EQUILIBRATION OF OXYGEN WITH MONOMERIC AND OCTAMERIC HEMERYTHRIN FROM THEMISTE-ZOSTERICOLA

Single relaxations for the equilibration of O2 with monomeric and octameric deoxy forms of hemerythrin from T. zostericola were observed at 25.degree. C using the temperature-jump technique. At 25.degree. C, pH 8.2 (Tris/H2SO4) and I = 0.10 M (Na2SO4), formation rate constants kon are 7.8 .cntdot. 107 M-1 .cntdot. s-1 and 7.5 .cntdot. 106 M-1 s-1, respectively. The procedure used does not give a precise measure (small intercepts) of dissociation rate constants, koff. These were determined instead by the stopped-flow method using dithionite to induce dissociation of the oxy protein. Values of koff for the monomer (3.1 .cntdot. 102 s-1) and octamer (82 s-1), in association with kon values, lead to equilibrium constants for the formation of oxyhemerythrin of 2.5 .cntdot. 105 M-1 and 0.9 .cntdot. 105 M-1, respectively, at 25.degree. C, pH 8.2 and I = 0.10 M (Na2SO4). These latter are in reasonable agreement with values (1.5 105 .cntdot. M-1 and 1.3 .cntdot. 105 M-1) determined spectrally on the equilibrated solutions. Using the octameric protein, it was shown that replacement of SO42- by ClO4- or Cl- ions (at a constant I = 0.10 M) led to .apprx. 2-fold enhancement of kon but had little effect on koff. The addition of Ca2+ or Mg2+ ions (0.01 M), with or without 0.50 M NaCl, also gives up to 4-fold increases in kon, but unchanged koff values. O2 pulse experiments on the octamer show no effect on koff of the degree of oxygenation of the protein. A comparison is made with similar data for Hb, myoglobin and hemocyanin.