MECHANISM OF ACTION OF THE LACTIC DEHYDROGENASE OF THE MAMMALIAN ERYTHROCYTE: II. THE MECHANISM OF SUBSTRATE INHIBITION

1958 ◽  
Vol 36 (1) ◽  
pp. 1085-1091 ◽  
Author(s):  
Paul Ottolenghi ◽  
Orville F. Denstedt
Keyword(s):  
Mechanism Of Action ◽  
Substrate Inhibition ◽  
Lactic Dehydrogenase ◽  
Mammalian Erythrocyte

The inhibition of lactic dehydrogenase produced by an excess of the substrate pyruvate has been shown to be of the noncompetitive type. Thus, two sites on the enzyme appear to be involved: one for the activation of pyruvate, and a second one which, when complexed with a molecule of pyruvate, renders the enzyme inactive.

MECHANISM OF ACTION OF THE LACTIC DEHYDROGENASE OF THE MAMMALIAN ERYTHROCYTE: II. THE MECHANISM OF SUBSTRATE INHIBITION

1958 ◽  
Vol 36 (10) ◽  
pp. 1085-1091 ◽  
Author(s):  
Paul Ottolenghi ◽  
Orville F. Denstedt
Keyword(s):  
Mechanism Of Action ◽  
Substrate Inhibition ◽  
Lactic Dehydrogenase ◽  
Mammalian Erythrocyte

The inhibition of lactic dehydrogenase produced by an excess of the substrate pyruvate has been shown to be of the noncompetitive type. Thus, two sites on the enzyme appear to be involved: one for the activation of pyruvate, and a second one which, when complexed with a molecule of pyruvate, renders the enzyme inactive.

MECHANISM OF ACTION OF THE LACTIC DEHYDROGENASE OF THE MAMMALIAN ERYTHROCYTE: I. INFLUENCE OF INHIBITORS

1958 ◽  
Vol 36 (10) ◽  
pp. 1075-1083 ◽  
Author(s):  
Paul Ottolenghi ◽  
Orville F. Denstedt
Keyword(s):  
Mechanism Of Action ◽  
Lactic Dehydrogenase ◽  
Noncompetitive Inhibition ◽  
Mammalian Erythrocyte ◽  
Lactate And Pyruvate

This paper pertains to the inhibition of lactic dehydrogenase by substances structurally related to its substrates, pyruvate and lactate. Oxalate, tartronate, and malonate inhibit the enzyme in a competitive manner towards lactate, and in a noncompetitive manner towards pyruvate. Phenoxyacetate causes simultaneous competitive and noncompetitive inhibition towards both lactate and pyruvate. The results obtained lead to the conclusion that, for their activation, pyruvate and lactate react with different sites on the enzyme surface.

MECHANISM OF ACTION OF THE LACTIC DEHYDROGENASE OF THE MAMMALIAN ERYTHROCYTE: I. INFLUENCE OF INHIBITORS

1958 ◽  
Vol 36 (1) ◽  
pp. 1075-1083 ◽  
Author(s):  
Paul Ottolenghi ◽  
Orville F. Denstedt
Keyword(s):  
Mechanism Of Action ◽  
Lactic Dehydrogenase ◽  
Noncompetitive Inhibition ◽  
Mammalian Erythrocyte ◽  
Lactate And Pyruvate

This paper pertains to the inhibition of lactic dehydrogenase by substances structurally related to its substrates, pyruvate and lactate. Oxalate, tartronate, and malonate inhibit the enzyme in a competitive manner towards lactate, and in a noncompetitive manner towards pyruvate. Phenoxyacetate causes simultaneous competitive and noncompetitive inhibition towards both lactate and pyruvate. The results obtained lead to the conclusion that, for their activation, pyruvate and lactate react with different sites on the enzyme surface.

Properties of Chlorophyllase from Capsicum annuum L. Fruits

2001 ◽  
Vol 56 (11-12) ◽  
pp. 1015-1021 ◽  
Author(s):  
Dámaso Hornero-Méndez ◽  
María Isabel Mínguez-Mosquera
Keyword(s):  
Substrate Specificity ◽  
Capsicum Annuum ◽  
Mechanism Of Action ◽  
Substrate Recognition ◽  
Substrate Inhibition ◽  
Inhibitory Effect ◽  
Hplc Separation ◽  
Substrate Complex ◽  
Enzyme Substrate

Abstract The in vitro properties of semi-purified chlorophyllase (chlorophyll-chlorophyllido hy­drolase, EC 3.1.1.14) from Capsicum annuum fruits have been studied. The enzym e showed an optimum of activity at pH 8.5 and 50 °C. Substrate specificity was studied for chlorophyll (Chi) a, Chi b, pheophytin (Phe) a and Phe b, with Km values of 10.70, 4.04, 2.67 and 6.37 μᴍ respectively. Substrate inhibition was found for Phe b at concentrations higher than 5 μᴍ. Chlorophyllase action on Chi a' and Chi b' was also studied but no hydrolysis was observed, suggesting that the mechanism of action depends on the configuration at C-132 in the chloro­ phyll molecule, with the enzyme acting only on compounds with R132 stereochemistry. The effect of various metals (Mg2+, Hg2+, Cu2+, Zn2+, Co , Fe2+ and Fe3+) was also investigated, and a general inhibitory effect was found, this being more marked for Hg2+ and Fe2+. Func­tional groups such as -SH and -S-S-seem ed to participate in the formation o f the enzyme-substrate complex. Chelating ion and the carbonyl group at C3 appeared to be important in substrate recognition by the enzyme. The method for measuring Chlase activity, including HPLC separation of substrate and product, has been optimized.

LACTIC DEHYDROGENASE OF THE MAMMALIAN ERYTHROCYTE: III. THE FORMATION OF A COMPLEX OF THE ENZYME WITH DPN AND LACTATE

1958 ◽  
Vol 36 (1) ◽  
pp. 1093-1098 ◽  
Author(s):  
Paul Ottolenghi ◽  
Orville F. Denstedt
Keyword(s):  
Ternary Complex ◽  
Kinetic Studies ◽  
Lactic Dehydrogenase ◽  
Binary Complex ◽  
Hydrogen Ions ◽  
Mammalian Erythrocyte

In the transfer of hydrogen ions and electrons from lactate to DPN by lactic dehydrogenase the lactate is capable of forming a complex only with the binary complex of enzyme and coenzyme to form an active ternary complex. The results of kinetic studies indicate that the site for the binding of lactate is on the DPN molecule itself.

scholarly journals Ο ρόλος των αμινοπεπτιδασών του ενδοπλασματικού δικτύου στην αντιγονοπαρουσίαση

2012 ◽  
Author(s):  
Ειρήνη Ευνουχίδου
Keyword(s):  
Immune Response ◽  
Molecular Mechanism ◽  
Mechanism Of Action ◽  
Molecular Model ◽  
Substrate Inhibition ◽  
Adaptive Immune Response ◽  
Sequence Specificity ◽  
Antigenic Peptides ◽  
Adaptive Immune ◽  
Molecular Mechanism Of Action

Endoplasmic reticulum aminopeptidases ERAP1/2 have emerged in the last years as key enzymes for the production of antigenic peptides that are presented by MHC class I molecules at the cell surface as part of the adaptive immune response. ERAP1 has unusual enzymatic properties that make it particularly suitable for this biological function. Specifically, it efficiently degrades peptides longer than 9 residues and shows preferences for the whole substrate sequence and not only for the N-terminus. Recently, coding Single Nucleotide Polymorphisms (SNPs) in ERAP1/2 were associated with predisposition to autoimmune diseases, such as ankylosing spondylitis, with cancer and with resistance to HIV infection. The hypothesis of this thesis is that the particular molecular mechanism of action of ERAP1/2 and the way in which this mechanism is affected by coding polymorphisms is the underlying reason for the association of ERAP1/2 with predisposition to human disease. To verify this hypothesis we investigated the molecular mechanism of action of ERAP1/2 aminopeptidases in the context of their polymorphic variability. We first developed a novel fluorigenic assay to analyze the activity of these enzymes. Biochemical analysis in combination with the recently solved ERAP1 crystal structure allowed us to propose a molecular model of function that can account for both substrate length and sequence specificity. In parallel, we set the groundwork for the development of small molecular weight compounds that modulate ERAP1 activity, by scanning a chemical library of pharmaceuticals and discovering lead compounds that either act as inhibitors or as activators of the enzyme.In order to understand the association of ERAP1/2 polymorphicity with changes in antigen presentation, we expressed different ERAP1/2 alleles and studied their ability to degrade different antigenic epitope precursors. Michaelis-Menten analysis showed that specific SNPs in ERAP1 affect the ΚΜ and kcat parameters of the enzyme and that particular allele-substrate combinations demonstrate substrate inhibition kinetics. A common ERAP2 allele was found to have a stronger effect on enzyme function and, as ERAP1 alleles, not only affects enzymatic activity but also alters specificity. The above results support the hypothesis that genetic variability in ERAP1/2 aminopeptidases can affect the generation of antigenic epitopes and may represent a previously unrecognized facet of adaptive immune response variability.

LACTIC DEHYDROGENASE OF THE MAMMALIAN ERYTHROCYTE: III. THE FORMATION OF A COMPLEX OF THE ENZYME WITH DPN AND LACTATE

1958 ◽  
Vol 36 (10) ◽  
pp. 1093-1098 ◽  
Author(s):  
Paul Ottolenghi ◽  
Orville F. Denstedt
Keyword(s):  
Ternary Complex ◽  
Kinetic Studies ◽  
Lactic Dehydrogenase ◽  
Binary Complex ◽  
Hydrogen Ions ◽  
Mammalian Erythrocyte

In the transfer of hydrogen ions and electrons from lactate to DPN by lactic dehydrogenase the lactate is capable of forming a complex only with the binary complex of enzyme and coenzyme to form an active ternary complex. The results of kinetic studies indicate that the site for the binding of lactate is on the DPN molecule itself.

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