Search PubMed⌕ Search

Biomedical subjects

M Brunori

Publications and source records attributed to M Brunori.

At least 343 records · Page 19Linked to original sources
PubMed1977

Oxygen "pulsed" cytochrome c oxidase: functional properties and catalytic relevance.

The kinetics of the reaction of cytochrome c with solubilized mammalian cytochrome c oxidase (ferrocytochrome c:oxygen oxidoreductase, EC 1.9.3.1) has been studied by a stopped-flow technique under two different experimental situations: (i) the completely oxidized enzyme (resting oxidase as obtained from the preparation) was mixed with reduced cytochrome c, and (ii) the completely reduced enzyme in the presence of reduced cytochrome c was exposed to a "pulse" of O2 (pulsed oxidase). Both sets of experiments were performed with either "limiting" or "excess" O2 (relative to oxidase), in the presence or absence of CO. Both the pre-steady-state events and the steady-state kinetics of cytochrome oxidase are found to be different in the two cases. This shows that the product of the reaction of fully reduced oxidase with O2 (pulsed oxidase) is functionally different from the oxidase as prepared (resting oxidase). These differences are interpreted with the assumption of a different rate of intramolecular electron transfer in the pulsed and resting oxidases. Implications of these experimental findings are discussed in the general framework of a tentative model for the catalytic cycle of the oxidase.

Animals↗
PubMed1976

Functional consequences of ligand-linked dissociation in hemoglobin from the sea cucumber Molpadia arenicola.

It has been established that Molpadia hemoglobin tends to dissociate into subunits as oxygen is bound. The kinetics and equilibria of carbon monoxide and ethylisocyanide binding reported here show a dependence on protein concentration that supports the conclusion that the aggregated hemoglobin has a lower ligand affinity than the dissociated subunits. This is true for the isolated D-chain as well as for the unfractionated hemolysate that contains four distinct polypeptide chains (A-D). This indicates that even homopolymers of Molpadia hemoglobin have lower ligand affinity than the dissociated subunits. At high protein concentration hemolysates of Molpadia hemoglobin show slight cooperativity. The time course of ligand binding to the deoxy D-chain also suggests cooperative interactions. The low affinity of the aggregated state may have a different molecular explanation than in human hemoglobin where tetramers of identical subunits (as in Hb H) show high oxygen affinity. The absence of tyrosine and histidine at the C-terminal of the Molpadia D-chains also suggests a different stabilization of the low affinity deoxy state. An additional functional difference between Molpadia hemoglobin and human hemoglobin is that organic phosphates do not alter the ligand affinity of the sea cucumber hemoglobin.

Animals↗
PubMed1976

Electron transfer between soluble and immobilized mammalian cytochrome c. Equilibrium and kinetic studies on immobilized cytochrome c.

Horse heart cytochrome c was covalently bound to Sepharose 4B and its redox properties were measured under various experimental conditions. The equilibrium constant for the electron exchange between the oxidized and the reduced form of cytochrome c when one of the two forms was in the semi-solid state and the other one in solution was close to 1. Matrix-bound ferrocytochrome c is very stable to autoxidation and is not oxidized by O2 even in the presence of mammalian cytochrome oxidase. Oxidation occurs if catalytic amounts of soluble cytochrome c are added to the reaction mixture. The rate of oxidation of matrix-bound ferrocytochrome c in the presence of cytochrome oxidase and catalytic amounts of soluble cytochrome c may be correlated with the rate of electron transfer between soluble and matrix-bound cytochrome c. This rate is more than two orders of magnitude lower than that reported for the homonuclear (between identical species) electron transfer in solution.

Animals↗
PubMed1976

The steady-state kinetics of the oxygenation of linoleic acid catalysed by soybean lipoxygenase.

The steady-state kinetics of the oxygenation of linoleic acid catalysed by soybean lipoxygenase-1 were studied. The results showed that lipoxygenase-1 is strongly inhibited by its substrate, linoleic acid. In the presence of the product of the reaction, 13-LS-hydroperoxy-linoleic acid, the substrate inhibition only affects the apparent affinity for O2 and is of a hyperbolic type. A kinetic scheme of the oxygenation reaction is presented, which postulates two substrate-binding sites on the enzyme, one for linoleic acid and one for O2, and a regulatory binding site, which can either bind the product or the fatty acid substrate. Since previous studies indicated that the product of the reaction influences the oxidation state of the iron present in protein, the steady-state kinetics of the native enzyme and of the enzyme pre-incubated with the product were compared. Pre-incubation of the enzyme with the product did not lead to altered steady-state kinetics of the reaction compared to those of the native enzyme.

Kinetics↗
PubMed1976

The virtual absence of antigenic cross-reactivity between functionally distinct trout hemoglobins.

The antigenic properties of the hemoglobins of two species of rainbow trout, Salmo irideus and Salmo gairdneri, have been compared. Each of these species possess two different classes of hemoglobins, the members of the first of which have no Bohr effect while those of the second exhibit a pronounced pH dependence of their ligand affinities. The hemoglobins from these two species are antigenically indistinguishable. However, between the two classes of hemoglobins there is almost no immunological cross-reactivity.

Animals↗
PubMed1976

Superoxide dismutase in red blood cells: method of assay and enzyme content in normal subjects and in patients with beta-thalassemia (major and intermedia).

The applicability to red blood cells of two widely used methods for spectrophotometric assay of superoxide dismutase activity has been tested, in view of the demand for routine screening of the level of this enzyme in various circumstances of hematological interest. The nitro blue tetrazolium reduction method was found to be inapplicable, even after removal of hemoglobin by chloroform-ethanol extraction. On the other hand, the epinephrine oxidation method gave reliable values after chloroform-ethanol extraction. The enzyme content in normal adults was found to be 6.2 +/- 1.4 X 10(15) gr. per red blood cell. No significant change in the enzyme content was found in 19 patients with beta-thalassemia major and 5 patients with beta-thalassemia intermedia, indicating no influence on the level of this enzyme by the increased flux of oxygen radicals which is to be expected in thalassemic red blood cells.

Adolescent↗
PubMed1975

Allosteric interactions in non-alpha chains isolated from normal human hemoglobin, fetal hemoglobin, and hemoglobin Abruzzo (beta143 (H21) His replaced by Arg).

Oxygen-linked effects of inositol hexaphosphate occur in heme-containing non-alpha chains isolated from normal human hemoglobin, fetal hemoglobin, and the abnormal human hemoglobin Abruzzo, beta143(H21) His leads to Arg. The occurrence of these effects implies that the chains undergo ligand-linked conformational changes. Inositol hexaphosphate lowers the oxygen affinity of isolated beta and gamma chains by differential binding to their deoxy conformations. Neither 2,3-diphosphoglycerate nor inorganic phosphate produces such an effect. In the case of Abruzzo beta chains, the binding of inorganic phosphate and 2,3-diphosphoglycerate is also oxygen-linked. Stripped beta chains isolated from hemoglobin Abruzzo have much higher oxygen affinity than beta chains isolated from HbA. Their higher oxygen affinity and enhanced allosteric interactions with phosphates account, in large part, for the abnormal functional behavior of the hemoglobin Abruzzo tetramer. In this hemoglobin variant the substitution of arginine for histidine at beta143 involves a residue known to interact with anionic allosteric effectors of hemoglobin. It is of interest that the effect of inositol hexaphosphate observed with isolated gamma chains is comparable to the effect observed with isolated beta chains, even though the gamma143 position is occupied by an uncharged serine residue.

Allosteric Regulation↗
PubMed1975

Hemoglobin Abruzzo (beta143 (H21) His replaced by Arg). Consequences of altering the 2,3-diphosphoglycerate binding site.

Hemoglobin Abruzzo is an abnormal human hemoglobin with a substitution at a residue known to be involved in the binding of 2,3-diphosphoglyceric acid. It has increased oxygen affinity and reduced heme-heme interaction in the absence of organic or inorganic phosphate cofactors. In inorganic phosphate buffers the Bohr effect and heme-heme interaction are normal, but the oxygen affinity remains higher than that of hemoglobin A. CO combination in inorganic phosphate is more strongly autocatalytic than in normal hemoglobin and a slower rate of oxygen dissociation is observed. Although many of the functional differences of this variant may be attributed to the high oxygen affinity of the mutant beta chains, the interactions between subunits are also affected by the histidine to arginine substitution at beta143. Stripped hemoglobin Abruzzo appears to be significantly more dissociated than hemoglobin A. Kinetic studies indicate that interaction with organic or inorganic phosphates decreases its subunit dissociation. In all of the functional properties examined, hemoglobin Abruzzo is more sensitive to the allosteric influence of organic and inorganic anions than is hemoglobin A.

Amino Acid Sequence↗