Hemoglobin rate constants

Unzai S et al Rate constants for O2 and CO binding to the alpha and beta subunits within the R and T states of human hemoglobin. J Biol Chem 1998 273 23150. 

Reversible electron transfer within Mg and Zn-substituted hemoglobin hybrids is initiated by flash photoproduction of the long-lived triplet state ( MP). According to Scheme I, the triplet return to the ground state involves two pathways, intrinsic triplet decay (with rate constant kp) and electron transfer quenching (with rate constant k,). 

The structural stability of mixed-metal hemoglobin hybrids also has allowed us to study low-temperature electron transfer in this system. We first reported the temperature dependence of triplet-state quenching within the [ (ZnP), Fe (H20)P] hybrids, which we attributed to the ZnP Fe P ET reaction [7d]. The rate constant dropped smoothly as the temperature was lowered from room temperature to 200 K. Below this temperature the rate constant remained roughly constant with a tunnelling rate constant of kt 9 s (Fig. 7). 

Metal-substituted hemoglobin hybrids, [MP, Fe " (H20)P] are particularly attractive for the study of long-range electron transfer within protein complexes. Both photoinitiated and thermally activated electron transfer can be studied by flash excitation of Zn- or Mg-substituted complexes. Direct spectroscopic observation of the charge-separated intermediate, [(MP), Fe " P], unambiguously demonstrates photoinitiated ET, and the time course of this ET process indicates the presence of thermal ET. Replacement of the coordinated H2O in the protein containing the ferric heme with anionic ligands (CN , F , Nj ) dramatically lowers the photoinitiated rate constant, k(, but has a relatively minor effect on the thermal rate, kg. 

Non-statistical successive binding of O2 and CO to the four heme centers of hemoglobin ( cooperativity ) has been thoroughly documented. It is difficult to test for a similar effect for NO since the equilibrium constants are very large ( 10 M ) and therefore difficult to measure accurately. It is found that the four successive formation rate constants for binding NO to hemoglobin are identical. In contrast, the rate constant for dissociation of the first NO from Hb(NO)4 is at least 80 times less than that for removal of NO from the singly bound entity Hb(NO). This demonstrates cooperativity for the system, and shows that it resides in the dissociation process. The thermodynamic implications of any kinetic data should therefore always be assessed. 

The system then returns to its initial state by back electron transfer from Fe(II)P to (ZnP)+. The decay of the TZnP triplet state is enhanced only when the p subunit contains Fe(III)P. When the j3 subunit contains Fe(II)P triplet, decay is unaffected. The rate constant of electron transfer, t, for [a(Zn)/J(Fe)] hybrid hemoglobin was found to be 102s 1 at room temperature [90]. The same value of kt was obtained for [a(Fe)/ (Zn)] hybrid hemoglobin [70]. 

The kinetics of the nonenzymatic glycosylation of hemoglobin to form hemoglobin Ale have been analyzed in detail by Higgins and Bunn (31). Assuming a bimolecular reaction for the condensation reaction and first-order kinetics for the Amadori rearrangement, they determined the rate constants for the glycosylation of the N-terminal valine of the 8-chain ... 

Zn cyt c/Fe(III) cyt b5] and [Zn hemoglobin/Fe(III) cyt bs] complexes have a similar structure. However, the rates of electron tunneling in these complexes are 102-fold different. This difference in rate constants may reflect direct differences in the conductivity towards electron tunneling of Hb vs cyt c. Clearly, subtle structural differences between these complexes are sufficient to cause large rate differences in the case of the tunneling mechanism. 

Temperature dependences of the rate for direct photoinduced electron transfer process and reverse charge recombination reaction were studied in some works. As a rule both processes were found to be temperature dependent. However for [p(MP), a(Fe(III)P hemoglobin hybrid (M = Zn(II), Mg(II)) the rate constants of both processes were found to be temperature independent in the temperature interval 273-293 K [285],... 

Temperature independent electron tunneling was observed also in Ref. [319], where the rate of electron transfer over large distance in mixed-metal hemoglobin hybrids [MP, FeIU(CN )P], where M = Zn or Mg, was measured in the temperature range from ambient to 100 K. The electron transfer from the triplet state of MP to Fera was not effected by the freezing of the cryosolvent, which may indicate that coupling of electron transfer to low-frequency solvent modes may be minimal. For both M, but especially for M = Mg, the rate constant of the back reaction is nearly temperature independent. 

The problem of base elimination in the CO kinetics was circumvented by measuring the kinetics of hemes with external 1- and 2-methylimidazole (20). The former is a strain-free system, and serves as a model for R-state hemoglobin, while the latter is strained (2-MeIm does not bind twice (21)), and serves as a model for T-state hemoglobin. The rate constants for addition of CO to the pentacoordinated heme-1-methylimidazole in CTAB buffer (Figure 5) are almost identical to those of R-state hemoglobin in Table I. The change from... 

The dependences, such as Eq. 2.47, are known as compensation effect, and coefficient (3C is denoted as isokinetic temperature at which all reactions of given series have the same rate constant. An example of compensation effect for for catalytic rate constant of the Sulfolobus solfataricus p-glycosidase reaction with different substrates is shown in Fig. 2.19. Similar relationships were reported for many other prosesses, involving the binding ligands to hemoglobin, the oxidation of alcohols by catalase, the hydroxylation of substrates by cytochrome c, etc. 

R. A. Alberty and G. G. Hammos, J. Phys. Chcm.y 62, 154 (1958), have shown that the very fast specific rate constants observed for this reversible reaction are compatible with a diffusion-controlled rate such that every collision of a fumarate ion with an active site leads to reaction. This is apparently true also for the combination of NO with iron hemoglobin and H2O2 with yeast peroxidase. 

In the synthetic iron porphyrin, O2 affinity mainly depends on the strengths of the <7-donation from the lone pair of O2 to the heme-iron dz orbital and 7r-back donation from the d/r orbital on the iron to the Jt orbital of 2. To evaluate the O2 affinity and/or O2 binding dynamics in myoglobin and hemoglobin, O2 - protein interaction is a further important factor. For example, the O2 dissociation rate constant for oxymyoglobin is relatively smaller than those of O2 complexes... 

Equilibrium and Rate Constants. Under physiological conditions, hemoglobin reacts with O2, CO, and NO to form oxyhemoglobin, Hb(02)4, carbonmonoxyhemoglobin, Hb(CO)4, and nitrosylhemoglobin, Hb(NO)4. Comparison of the relative equilibrium constants, A f rei, for formation of the fully loaded complexes ... 

The decontamination efficacy of radioprotective agent aminoethylisothiourea and cystamine on rat skin against SM was comparable to standard decontaminants, such as alco-holate, clay, and Fuller s earth (Knezevic and Tadic, 1996). A mixture containing bovine hemoglobin, gelatin, and poi (a Hawaiian foodstuff) on the hydrolytic kinetics of the mustard simulants, 2-chloroethyl ethyl sulfide, 2-chloro-ethyl methyl sulfide and 2-bromoethyl phenyl sulfide, is reported. The kinetic mechanisms and rate constants were dependent upon the mixtures concentration and viscosity (Cemy and Cemy, 1997). 

Important reactions of NO are binding to transition metals like heme iron as in hemoglobin to produce nitroso-hemoglobin, addition to thiols to produce nitrosothiols as given in Eqs. (8) and (9). The rate constants for the two reactions are reported by several groups and found to be in the range (3-5) x 10 and (1-3) x 10 M" s" ... 

In contrast, amyl nitrite can cause methemoglobinemia when abused, since it has a 2000 times higher affinity for hemoglobin (58). In vitro both amyl nitrite and glyceryl trinitrate reacted immediately with oxyhemoglobin to effect oxidation to methemoglobin, while for sodium nitrite there was a lag phase before the reaction occurred. The affinity rate constants were ... 

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