Oxygen equilibrium properties of nickel(II)-iron(II) hybrid hemoglobins cross-linked between 82 beta 1 and 82 beta 2 lysyl residues by bis(3,5-dibromosalicyl)fumarate: determination of the first two-step microscopic Adair constants for human hemoglobin.
We have previously reported that cross-linked asymmetric Ni(II)-Fe(II) hybrid hemoglobin, XL[alpha (Fe) beta (Fe)][alpha (Ni) beta (Ni)], in which the alpha 1 beta 1 dimer containing ferrous protoporphyrin IX and the adjacent alpha 2 beta 2 dimer containing nickel(II) protoporphyrin IX were cross-linked between Lys-82 beta 1 and Lys-82 beta 2 by reaction with bis(3,5-dibromosalicyl)fumarate, represents an adequate model for determination of the alpha 1 beta 1 oxygenation properties of native hemoglobin [Shibayama, N., Imai, K., Morimoto, H., & Saigo, S. (1993) Biochemistry 32, 8792-8798]. To extend the approach using cross-linked Ni(II)-Fe(II) hybrids to all possible pathways for initial-half oxygenation of hemoglobin, we have prepared three other types of cross-linked Ni(II)-Fe(II) hybrids, carrying nickel(II) protoporphyrin IX in two subunits and ferrous protoporphyrin IX in the other two subunits, and have determined the two-step oxygen equilibrium curves of the ferrous subunits within these cross-linked hybrids. For the first step of oxygenation, the alpha subunit shows about 3-fold higher affinity than the beta subunit at all pH values examined, indicative of a significant functional heterogeneity of the subunits in deoxyhemoglobin. For the second step of oxygenation, the cooperativity represented by the Hill coefficient (nmax) increases in the order of beta 1 beta 2 (nmax = 1.36), alpha 1 beta 1 (nmax = 1.41), alpha 1 beta 2 (nmax = 1.64), and alpha 1 alpha 2 (nmax = 1.72) at pH 7.4 in the presence of 0.1 M Cl- at 25 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)
Shibayama
N
N
Department of Physics, Jichi Medical School, Tochigi, Japan.
Imai
K
K
Morimoto
H
H
Saigo
S
S
D001241:Aspirin / Q000031:analogs & derivatives* / Q000737:chemistry; D003432:Cross-Linking Reagents / Q000737:chemistry; D006454:Hemoglobins / Q000737:chemistry*; D006801:Humans; D007501:Iron / Q000737:chemistry*; D008239:Lysine / Q000737:chemistry; D009532:Nickel / Q000737:chemistry*; D010100:Oxygen / Q000737:chemistry*; D055503:Protein Multimerization
D003432:Cross-Linking Reagents; D006454:Hemoglobins; C053220:nickel-iron hybrid hemoglobin; C021630:bis(3,5-dibromosalicyl)fumarate; D009532:Nickel; D007501:Iron; D008239:Lysine; D001241:Aspirin; D010100:Oxygen