Association of methanol and ethanol with heme proteins

As, Brill; Bw, Castleman; Me, McKnight

doi:10.1021/bi00656a010

Cited by 32 publications

(16 citation statements)

References 20 publications

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“…The subsequent spin-state transition (500 ps/div) proceeds with a rate constant close to 1 ms-' and is clearly distinguished from the reduction process. The concentration of methemoglobin was 46.6 pM and the reduction degree 0.02 cording to Brill et al [13], just coincides with a maximum of the stability of the protein structure which in this case is at least as stable as in a methanol-free buffer. According to Hollocher [14], methanol in aqueous solutions strongly enhances the formation of ferric hemochromes.…”

Section: Pulse Radiolysis Of Methemoglohin In Aqueous-metlzanol Solutsupporting

confidence: 74%

The Spin‐State Transition of the Hemochrome Non‐equilibrium Conformation in Partially Reduced Human Methemoglobin

Raap

Leeuwen

Rollema

et al. 1978

European Journal of Biochemistry

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The effect of external parameters on the relaxation process of the hemochrome-type non-equilibrium conformation in partially reduced methemoglobin has been investigated. The relaxation of the intermediate ferrous low-spin state to the high-spin equilibrium conformation of hemoglobin appears to be facilitated particularly by protons and phosphate ions. In addition to studying the spin-state transition in aquomethemoglobin we have also studied it in complexes of the heme group in methemoglobin with fluoride, azide and cyanide anions.The observation of a spin-state transition of the iron atom after reduction of the ferric heme group in methemoglobin by solvated electrons prompted us to examine this process in more detail. In a preliminary communication we reported that the visible absorbance spectrum observed immediately after the actual reduction process of the heme group revealed a low-spin ferrous heme spectrum which is characteristic for a hemochrorne state [l] T(t) state, the tense quaternary (tertiary) struc-~LII-C: I<" \(ate. a n activated state of the quaternary R structure; d; I and d: 2 , thc low-spin and high-spin state of the ferric ion; d: and d$, the low-spin and high-spin state of the Ferrous ion. Bistris, 2,2~bis(hydroxymethy1)-2,2',2"-nitrilotrietIianol. globin [3]. Previously we suggested [l] that the distal hi\tidyl residue is probably the primary electron acceptor of the solvated electron; subsequently it acts as an electron donor during the electron transfer process ; in the course of the process this residue becomes bound to the iron atom of the heme group and remains bound until the spin-state transition removes it again. This suggests that the spin-state transition will certainly be accompanied by structural changes within the protein conformation.From our studies on the binding kinetics of carbon monoxide [4,5] and oxygen [6] for partially reduced methemoglobin and for fully reduced valency hybrids of hemoglobin (consisting of two identical oxidized subunits and two identical CO-liganded subunits), we obtained information about the quaternary structure of the hemoglobin intermediates. In particular the study of the binding of carbon monoxide to fully reduced valency hybrids suggested the existence of an activated transient R" state which rapidly decays into the quaternary R state or T state. The non-equilibrium R* state has tentatively been ascribed by us to the intermediate hernochrome state [ 5 ] . In order to elucidate the physico-chemical properties of the hernochrome state we started an extensive investigation of the influence that the solute conditions and the ligand bound to the ferric heme group have on the decay rate of this non-equilibrium conformation.

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Section: Pulse Radiolysis Of Methemoglohin In Aqueous-metlzanol Solutsupporting

confidence: 74%

The Spin‐State Transition of the Hemochrome Non‐equilibrium Conformation in Partially Reduced Human Methemoglobin

Raap

Leeuwen

Rollema

et al. 1978

European Journal of Biochemistry

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“…The biphasical behaviour of the oxygenation process is independent of the methanol concentration in the range of 20 to 1000 mM. Therefore the interaction of methanol with methemoglobin leading to a methemoglobin-methanol complex with a dissociation constant of 45 mM [34] introduces no heterogeneity with respect to the oxygenation of Hb3+. The average value of the faster fractionf, equal to 0.63 0.07 at pH = 7, persisted after large variation in this external parameter.…”

Section: Discussionmentioning

confidence: 95%

“…The addition of 100 mM methanol to the solutions of methemoglobin is sufficient to scavenge the hydroxyl radical shortly after the submicrosecond electron-pulse. However, it has been found by Brill et al [34] that a methemoglobin-methanol complex is formed, with a dissociation constant of about 45 mM. To find out whether the radical scavenger influences the relative contribution of both phases during the oxygenation process of Hb3 +, two extreme concentrations of methanol were taken ( Table 1).…”

Section: Heterogeneous Oxygeriation 0fhb3+mentioning

confidence: 99%

Heterogeneity in the Kinetics of Oxygen Binding to Partially Reduced Human Methemoglobin

Raap

Leeuwen

Eck-Schouten

et al. 1977

European Journal of Biochemistry

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The pulse-radiolysis technique has been introduced because it permits a rapid reduction (in a few microseconds) of one heme group of the methemoglobin tetramer by hydrated electrons. The kinetics of the binding of oxygen to this particular valence intermediate (Hb3+) with one reduced a or p subunit has been studied. It appears that the hydrated electrons preferentially reduce one type of subunit of methemoglobin at acid and neutral pH-values as is shown by the biphasic behaviour of Hb3+ on oxygenation. The second-order on-rate constants measured for the binding of oxygen to Hb3+ are 14 k 3 mM-' ms-' and 56 f 9 mM-' ms-I, respectively. The relative contribution of the faster fraction is about 0.63 f 0.08 of the total oxygenation process.A comparison of the kinetic absorbance difference spectrum for the reduction of methemoglobin with the static difference spectrum of deoxyhemoglobin and methemoglobin in the Soret-region revealed a decreased absorbance of the unliganded subunit of Hb3+ at 430 nm. This fact suggests that Hb3+ is in the relaxed quaternary conformation, which is in agreement with the observed on-rate constants.The intrinsic association and on-rate constants for the successive binding of oxygen to hemoglobin tetramers increase at each ligation step. This property of the hemoglobin molecule leads to the well-known sigmoid shape of the saturation curve for oxygen binding, indicating a positive mode of interaction between the subunits of the hemoglobin tetramer [1,2]. Monod et al. [3] proposed that the existence of a low affinity state for unliganded hemoglobin (T-state) and for a high affinity state of liganded hemoglobin (Rstate) can explain the positive mode of interaction. These two states of the MWC-model correspond to the fully liganded and unliganded X-ray structures observed by Perutz et al. [4].Recently it has been suggested by several investigators [5 -81 that the cooperative interactions of hemoglobin are not determined exclusively by the two quaternary conformations. The observed differences Abbreviations and Symbols. MetHb, methemoglobin; Hb4', Hb3+, Hb", €Ib+ : the superscripts refer to the number of oxidized subunits of the hernoglobin tetramer; Hb, deoxyhemoglobin; Hb*, rapidly reacting hemoglobin (R-state); HbOz, oxyhemoglobin; Bistris, [bis(2-hydroxyethyl)amino]tris (hydroxymethy1)methane.in the ligation and the redox reaction properties of the a and B subunits indicate that the predominance of one type of subunit may introduce an apparent slight negative cooperativity into the hemoglobin tetramer. It is evident that the non-equivalence of the subunits will be observed most favourably when the hemoglobin molecules are in the same quaternary conformation. The purpose of this paper is to establish the contribution of the a-p chain heterogeneity to the partial reduction of methemoglobin to Hb3+ and the kinetics of oxygen binding to this species.It is more difficult to determine the intrinsic reaction rate constants for the combination of oxygen with hemoglobin than for carbon monoxide to ...

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“…The overall relaxation rate is notably slower in myoglobin solutions, however. The dissociation constant for the methanol complex with myoglobin is 170 mM at 25 OC (Brill et al, 1976). The rate of exchange of a methanol ligand from the heme in myoglobin (calculated by using eq 3) is 9.5 X lo2 s-l at 25 O C , which is slower than for leghemoglobin (Table 111).…”

Section: Effects Of Acetone and Methanol On Leghemoglobinmentioning

confidence: 95%

“…Our own unpublished experiments show many heme proteins to be sensitive to the presence of even moderate concentrations of organic solvents. Binding of methanol and ethanol to ferric myoglobin and ferric hemoglobin has recently been reported (Brill et al, 1976). Reactions of peroxidase with hydrogen peroxide or cyanide are inhibited by ethanol, and it has been suggested that this alcohol binds to the sixth coordination position of the heme (Dunford & Hewson, 1977).…”

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confidence: 99%

Measurement of heme accessibility in soybean ferric leghemoglobin a and its complexes by proton magnetic relaxation

Ollis¹,

Wright²,

Pope³

et al. 1981

Biochemistry

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The accessibility of the heme and solvent-exchange dynamics in solutions of soybean ferric leghemoglobin a and several of its complexes have been determined by measurement of the relaxation rates of water, acetone, and methanol protons. Acetone and methanol were used at low concentrations as probes of outer-sphere and inner-sphere exchange processes, respectively. Water proton relaxation in solutions of ferric leghemoglobin is exchanged limited. The rate of exchange of water molecules or protons from the environment of the heme is greater than that for ferric myoglobin and ferric hemoglobin. Methanol binds to ferric leghemoglobin in a homogeneous first-order reaction and in competition with fluoride, a known iron ligand. Both optical and NMR measurements indicate binding of methanol to the sixth coordination position of the iron atom. Methanol proton relaxation by both ferric leghemoglobin and whale ferric myoglobin is w t e r proton relaxation enhancement (PRE)' measurements in solutions of ferric heme proteins have been widely used to probe the immediate environment of the heme and its accessibility to solvent molecules. Interpretation of the data is complex, however, since relaxation of protons of the bulk solvent may arise from inner-sphere (exchange of water molecules or protons from the coordination sphere of the ferric ion) or outer-sphere (exchange of noncoordinated water molecules or protons from the vicinity of the metal ion) exchange processes. A stereochemical marker method, in which the relaxation of the aliphatic protons of alcohols in the heme protein solutions is measured, has been proposed to distinguish between these relaxation mechanisms (Vuk-PavloviE et al., 1974). However, at the high concentrations of alcohols used by these workers there is considerable risk of conformational perturbation of the protein. Our own unpublished experiments show many heme proteins to be sensitive to the presence of even moderate concentrations of organic solvents. Binding of methanol and ethanol to ferric myoglobin and ferric hemoglobin has recently been reported (Brill et al., 1976). Reactions of peroxidase with hydrogen peroxide or cyanide are inhibited by ethanol, and it has been suggested that this alcohol binds to the sixth coordination position of the heme (Dunford & Hewson, 1977). In view of the evidence for interactions between alcohols and heme proteins, we have used a modification of the stereochemical marker method (Vuk-PavloviE et al., 1974) together with conventional water PRE measurements in studies of proton relaxation in solutions of heme proteins. The present method differs from the stereochemical marker method in that only low, nonperturbing concentrations of the

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Association of methanol and ethanol with heme proteins

Cited by 32 publications

References 20 publications

The Spin‐State Transition of the Hemochrome Non‐equilibrium Conformation in Partially Reduced Human Methemoglobin

The Spin‐State Transition of the Hemochrome Non‐equilibrium Conformation in Partially Reduced Human Methemoglobin

Heterogeneity in the Kinetics of Oxygen Binding to Partially Reduced Human Methemoglobin

Measurement of heme accessibility in soybean ferric leghemoglobin a and its complexes by proton magnetic relaxation

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