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The mechanism of haem catabolism. A study of haem breakdown in spleen microsomal fraction and in a model system by 18O labelling and metal substitution.

ABSTRACT: The mechanism of bile-pigment formation from haem breakdown was studied by using 18O labelling of the molecular oxygen required for macrocyclic ring cleavage. For haem degradation by the spleen microsomal haem oxygenase system, mass spectrometry of the product bilirubin revealed that cleavage occurred by the Two-Molecule Mechanism, i.e. the terminal lactam oxygen atoms in bilirubin were derived from two different oxygen molecules. Similarly, degradation of myoglobin by coupled oxidation with ascorbate and oxygen proceeded via the Two-Molecule Mechanism. Cobalt and manganese complexes of protoporphyrin IX were not degraded by either the haem oxygenase system or the coupled oxidation system. This result suggests that the iron atom possesses unique properties in facilitating porphyrin breakdown.

SUBMITTER: King RF

PROVIDER: S-EPMC1185890 | biostudies-other | 1978 Jul

REPOSITORIES: biostudies-other

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The mechanism of haem catabolism. Bilirubin formation in living rats by [18O]oxygen labelling.

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| S-EPMC1183896 | biostudies-other

An 18O double-labelling study of haemoglobin catabolism in the rat.

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Stable Isotope Phosphate Labelling of Diverse Metabolites is Enabled by a Family of 18O‐Phosphoramidites**

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A study of the mechanism of quercetin oxygenation by 18O labelling. A comparison of the mechanism with that of haem degradation.

Project description:The mechanism of quercetin oxygenation, which is formally analogous to haem degradation, was studied by using 18O labelling. In both the enzymic oxygenation (catalysed by quercetinase) and the non-enzymic reaction (base-catalysed), both oxygen atoms incorporated into product were derived from a single oxygen molecule. Quercetin oxygenation therefore occurs by a classical dioxygenase mechanism and is not an appropriate model for study of the mechanism of haem catabolism.

| S-EPMC1158469 | biostudies-other

Electron-transport pathway of the NADH-dependent haem oxygenase system of rat liver microsomal fraction induced by cobalt chloride.

Project description:The hepatic microsomal haem oxygenase activity of rats treated with CoCl2 was studied kinetically by measuring biliverdin, the immediate product of the reaction. Biliverdin was extracted with diethyl ether/ethanol mixture, and was determined by the difference between A690 and A800. The apparent Km value for NADPH (at 50 microM-haematin) was about 0.2 microM when an NADPH-generating system was used, whereas that for NADH was about 630 microM. Essentially the same Vmax. values were obtained for both the NADH- and NADPH-dependent haem oxygenase reactions. No synergism was observed with NADH and NADPH. The NADH-dependent reaction was competitively inhibited by NADP+, with a Ki of about 10 microM. The inhibitoin of the NADH-dependent reaction by the antibody against rat liver microsomal NADPH-cytochrome c reductase was essentially complete, with a pattern similar to that of the NADPH-dependent reaction. The immunochemical experiment and the comparison of the kinetic values with the reported data on isolated NADH-cytochrome b5 reductase and NADPH--cytochrome c reductase indicated the involvement of the latter enzyme in NADH-dependent haem oxygenation by microsomal fraction in situ.

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Metal substitution of tetanus neurotoxin.

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Accelerated hepatic haem catabolism in the selenium-deficient rat.

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| S-EPMC1184121 | biostudies-other

Proteome turnover of Arabidopsis root microsomal fraction

Project description:8-day-old seedlings were transferred onto media containing 15N-salts and were collected at 0, 4, 8, 16, 24, 32, 40, and 48 hours after transfer.

2015-08-12 | MSV000079233 | MassIVE

UDP-glucosyltransferase activity of housefly microsomal fraction.

Project description:Housefly UDP-glucosyltransferase activity towards p-nitrophenol was demonstrated in a system in vitro. The activity is localized in the microsomal fraction, requires UDP-glucose, is slightly stimulated by Mg2+ and is activated optimally over a wide range of detergent concentration. Phenobarbital increases the enzyme(s) activity about 3-fold, with p-nitrophenol as substrate, which differs from the corresponding mammalian glucuronyltransferase.

| S-EPMC1186444 | biostudies-other

Nickel quercetinase, a "promiscuous" metalloenzyme: metal incorporation and metal ligand substitution studies.

Project description:BackgroundQuercetinases are metal-dependent dioxygenases of the cupin superfamily. While fungal quercetinases are copper proteins, recombinant Streptomyces quercetinase (QueD) was previously described to be capable of incorporating Ni(2+) and some other divalent metal ions. This raises the questions of which factors determine metal selection, and which metal ion is physiologically relevant.ResultsMetal occupancies of heterologously produced QueD proteins followed the order Ni > Co > Fe > Mn. Iron, in contrast to the other metals, does not support catalytic activity. QueD isolated from the wild-type Streptomyces sp. strain FLA contained mainly nickel and zinc. In vitro synthesis of QueD in a cell-free transcription-translation system yielded catalytically active protein when Ni(2+) was present, and comparison of the circular dichroism spectra of in vitro produced proteins suggested that Ni(2+) ions support correct folding. Replacement of individual amino acids of the 3His/1Glu metal binding motif by alanine drastically reduced or abolished quercetinase activity and affected its structural integrity. Only substitution of the glutamate ligand (E76) by histidine resulted in Ni- and Co-QueD variants that retained the native fold and showed residual catalytic activity.ConclusionsHeterologous formation of catalytically active, native QueD holoenzyme requires Ni(2+), Co(2+) or Mn(2+), i.e., metal ions that prefer an octahedral coordination geometry, and an intact 3His/1Glu motif or a 4His environment of the metal. The observed metal occupancies suggest that metal incorporation into QueD is governed by the relative stability of the resulting metal complexes, rather than by metal abundance. Ni(2+) most likely is the physiologically relevant cofactor of QueD of Streptomyces sp. FLA.

| S-EPMC4416304 | biostudies-literature

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