Mixed substrate experiments with human brain monoamine oxidase.
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Amines↗
Biomedical subjects
M D Houslay
Publications and source records attributed to M D Houslay.
A kinetic evaluation of monoamine oxidase activity in rat liver mitochondrial outer membranes.
1. A preparation of mitochondrial outer membranes from rat liver can be shown to contain two kinetically distinct monoamine oxidase activities. These activities are distinguishable by their different sensitivities to the irreversible inhibitor clorgyline, and by the effect of the reversible inhibitors benzyl cyanide and 4-cyanophenol. 2. The substrate specificities of the preparation and the two enzyme species have been elucidated.
Acetonitriles↗
Allotopic properties of human brain monoamine oxidase.
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Alkynes↗
The nature of the electrophoretically separable multiple forms of rat liver monoamine oxidase.
1. Treatment of a partly purified preparation of rat liver monoamine oxidase with the chaotropic agent sodium perchlorate caused the enzyme to migrate as a single band of activity of polyacrylamide-gel electrophoresis, whereas the untreated enzyme separated into a number of bands. 2. Treatment with the chaotropic agent caused no loss of enzyme activity towards benzylamine, dopamine or tyramine. 3. The activities of the untreated preparation towards different substrates were inhibited to different extents by heat treatment and by some inhibitors. No such differences could be detected after the enzyme preparation had been treated with sodium perchlorate. 4. Lipid material, which could be separated by gel filtration, was liberated from the enzyme preparation by sodium perchlorate treatment. 5. The molecular weight of the treated enzyme was found to be 380000+/-38000. 6. Perchlorate treatment altered the solubility of the enzyme. 7. A continuous assay method for monoamine oxidase is described.
Animals↗
The reaction pathway of membrane-bound rat liver mitochondrial monoamine oxidase.
1. A preparation of a partly purified mitochondrial outer-membrane fraction suitable for kinetic investigations of monoamine oxidase is described. 2. An apparatus suitable for varying the O(2) concentration in a spectrophotometer cuvette is described. 3. The reaction catalysed by the membrane-bound enzyme is shown to proceed by a double-displacement (Ping Pong) mechanism, and a formal mechanism is proposed. 4. KCN, NaN(3), benzyl cyanide and 4-cyanophenol are shown to be reversible inhibitors of the enzyme. 5. The non-linear reciprocal plot obtained with impure preparations of benzylamine, which is typical of high substrate inhibition, is shown to be due to aldehyde contamination of the substrate.
Amines↗
Activation of two signal-transduction systems in hepatocytes by glucagon.
The ability of glucagon to stimulate glycogen breakdown in liver played a key part in the classic identification of cyclic AMP and hormonally stimulated adenylate cyclase. But several observations indicate that glucagon can exert effects independent of elevating intracellular cAMP concentrations. These effects are probably mediated by an elevation of the intracellular concentration of free Ca2+ although the mechanism by which this occurs is unknown. We show here that glucagon, at the low concentrations found physiologically, causes both a breakdown of inositol phospholipids and the production of inositol phosphates. Indeed, we show that the glucagon analogue, (1-N-alpha-trinitrophenylhistidine,12-homoarginine)glucagon (TH-glucagon), which does not activate adenylate cyclase or cause any increase in cAMP in hepatocytes yet can fully stimulate glycogenolysis, gluconeogenesis and urea synthesis, stimulates the production of inositol phosphates. This stimulation of inositol phospholipid metabolism by low concentrations of glucagon provides a mechanism whereby glucagon can exert cAMP-independent actions on target cells. We suggest that hepatocytes possess two distinct receptors for glucagon, a GR-1 receptor coupled to stimulate inositol phospholipid breakdown and a GR-2 receptor coupled to stimulate adenylate cyclase activity.
Adenylyl Cyclases↗
Normal p21N-ras couples bombesin and other growth factor receptors to inositol phosphate production.
Many receptors, in response to ligand activation, trigger inositol phospholipid breakdown, which leads to rapid intracellular responses. The sustained activation of this pathway is believed to be at least one of the factors involved in the stimulation of cell growth and there has been much speculation that certain oncogenes use this pathway to effect uncontrolled cellular proliferation. It has been suggested, by analogy with the receptor-mediated control of adenylate cyclase, that the receptor stimulation of inositol phospholipid metabolism is mediated through a guanine nucleotide regulatory protein (G-protein) called Gp (or Np). Although such a species has not been identified, there is now strong experimental evidence that this process is mediated by a G-protein distinct from the stimulatory and inhibitory G-proteins (Gs and Gi, respectively). The ras genes code for a plasma membrane protein, p21, whose only known biochemical property is a high-affinity GTPase activity. We show here that the expression of normal p21N-ras in NIH 3T3 fibroblasts leads to the coupling of certain growth factor receptors to stimulated inositol phosphate production. We propose that the N-ras proto-oncogene encodes a protein which couples the receptors for certain growth factors to the stimulation of phospholipase C. Thus, N-ras p21 may be the putative Gp or a functionally related protein.
Animals↗
Abolition of the expression of inhibitory guanine nucleotide regulatory protein Gi activity in diabetes.
Many cell-surface receptors for hormones appear to exert their effects on target cells by interacting with specific guanine nucleotide binding regulatory proteins (G-proteins) which couple receptors to their second-messenger signal generation systems. A common intracellular second messenger, which is used by many hormones, is cyclic AMP. This is produced by adenylate cyclase, whose activity is controlled by two G-proteins, Gs which mediates stimulatory effects and Gi inhibitory effects on adenylate cyclase activity. In liver, the hormone glucagon increases intracellular cAMP concentrations by activating adenylate cyclase by a Gs-mediated process. This effect of glucagon is antagonised by the hormone insulin, although the molecular mechanism by which insulin elicits its actions is obscure. However, insulin receptors exhibit a tyrosyl kinase activity and appear to interact with G-proteins, perhaps by causing phosphorylation of them. In type I diabetes, circulating insulin levels are abnormally low, giving rise to gross perturbations of metabolism as well as to a variety of complications such as ionic disturbances, neuropathies of the nervous system, respiratory and cardiovascular aberrations and predisposition to infection. We show here that experimentally-induced type I diabetes leads to the loss of expression of Gi in rat liver. As it has been suggested that Gi may couple receptors to K+-channels as well as mediating the inhibition of adenylate cyclase, aberrations in the control of expression of this key regulatory protein in type I diabetes may be expected to lead to pleiotropic effects.
Adenosine Diphosphate Ribose↗
Localization of the gene for the human serotonin 5-HT(2B) receptor to chromosome 2.
A 483 bp Bgl II/Sal I repeat free fragment was isolated from the vector containing the human serotonin 5-HT(2B) receptor. This fragment contained part of the coding sequence located within the last exon and the entire 3'-non-translated region. It was used to probe, by Southern blots, Pst I digested DNA from a series of 20 rodent-human somatic cell hybrid lines, each of which contained different complements of human chromosomes. This 5-HT(2B) receptor probe showed hybridization to a 2-7 kb Pst I fragment which revealed 100 percent concordance with location of the gene for this 5-HT receptor variant at chromosome 2.
Animals↗