Modification of arginine residues of pea lactate dehydrogenase by phenylglyoxal

1990 ◽  
Vol 55 (5) ◽  
pp. 1380-1388
Author(s):  
Jana Barthová ◽  
Jana Kunová ◽  
Sylva Leblová
Keyword(s):  
Lactate Dehydrogenase ◽  
Dissociation Constants ◽  
Ionic Bond ◽  
Complete Inactivation ◽  
Pea Seedlings ◽  
Arginine Residues ◽  
Michaelis Constants ◽  
Lactate And Pyruvate ◽  
Sepharose 4B

Electrophoretically homogeneous lactate dehydrogenase was isolated from germinating pea seedlings by chromatography on AMP-Sepharose 4B. The amino acid composition of the enzyme was determined as well as the values of the Michaelis constants for four substrates and of the dissociation constants for the binary enzyme-coenzyme complexes. The treatment of the enzyme with phenylglyoxal resulted in the modification of nine arginine residues in its subunit. The modification was paralleled by a complete inactivation of the enzyme. The role of the arginine residues in the active center probably involves the binding of substrates, lactate and pyruvate, to the apoenzyme by an ionic bond.

Interaction of lactate dehydrogenase isoenzymes with ligands

1988 ◽  
Vol 53 (8) ◽  
pp. 1857-1861 ◽  
Author(s):  
Jana Barthová ◽  
Jana Kučerová ◽  
Sylva Leblová
Keyword(s):  
Lactate Dehydrogenase ◽  
Enzyme Inhibitor ◽  
Dissociation Constants ◽  
Physiological Role ◽  
Competitive Inhibitor ◽  
Kinetic Measurements ◽  
Cibacron Blue ◽  
Michaelis Constants ◽  
Sepharose 4B ◽  
Lactate Dehydrogenase Isoenzymes

Isoenzymes of bovine lactate dehydrogenase (H4, H3M, and H2M2) were prepared by affinity chromatography on a 5'-AMP-Sepharose 4B column. The interaction of isoenzymes with two ligands, coenzyme NADH and the competitive inhibitor Cibacron Blue F3GA was followed by means of kinetic measurements and by affinity electrophoresis. The Michaelis constants of NADH were compared with the inhibition constants of Cibacron Blue and dissociation constants of enzyme-inhibitor complexes. It was found that the M subunit of lactate dehydrogenase exhibits always higher affinity both to NADH and Cibacron Blue in comparison to the H subunit. This finding corresponds to the physiological role of lactate dehydrogenase isoenzymes.

Isolation of plant lactate dehydrogenase by affinity chromatography and role of histidine in the molecule of the enzyme

1980 ◽  
Vol 45 (5) ◽  
pp. 1608-1615 ◽  
Author(s):  
Jana Barthová ◽  
Pavla Plachá ◽  
Sylva Leblová
Keyword(s):  
Lactate Dehydrogenase ◽  
Affinity Chromatography ◽  
Inactivation Rate ◽  
Molar Ratio ◽  
Coenzyme Binding ◽  
Soybean Seedlings ◽  
Diethyl Pyrocarbonate ◽  
Binary Complexes ◽  
Sepharose 4B

Lactate dehydrogenase (EC 1.1.1.27) was isolated from soybean seedlings (Glycine max. L.) by affinity chromatography on an AMP-Sepharose 4B column. The enzyme obtained was inactivated by treatment with diethyl pyrocarbonate; the inactivation rate was proportional to the molar ratio of the enzyme to the reagent. The plot of the inactivation rate versus pH shows that of all the functional groups of the protein the imidazole groups of histidine only were modified by diethyl pyrocarbonate. By this procedure 20 histidine residues were ethoxyformylated in the molecule of soybean lactate dehydrogenase yet 8 only, i.e. two in every subunit were essential for thae activity of the enzyme. A comparison of the effect of diethyl pyrocarbonate on the lactate dehydrogenase apoenzyme with its effect on the binary complexes of the enzyme with coenzymes or on ternary complexes with its both substrates permits the conclusion that histidine is involved not only in the proton transfer during the redox reaction but also in the coenzyme-binding site.

scholarly journals Mr 46,000 mannose 6-phosphate receptor. The role of histidine and arginine residues for binding of ligand.

1991 ◽  
Vol 266 (5) ◽  
pp. 2917-2923 ◽  
Author(s):  
M Wendland ◽  
A Waheed ◽  
K von Figura ◽  
R Pohlmann
Keyword(s):  
Arginine Residues ◽  
Mannose 6 Phosphate

scholarly journals Species-Specific Regulation of TRPM2 by PI(4,5)P2 via the Membrane Interfacial Cavity

2021 ◽  
Vol 22 (9) ◽  
pp. 4637
Author(s):  
Daniel Barth ◽  
Andreas Lückhoff ◽  
Frank J. P. Kühn
Keyword(s):  
Amino Acid Residues ◽  
Hek 293 ◽  
Complete Inactivation ◽  
293 Cells ◽  
Activation Mechanisms ◽  
Specific Regulation ◽  
Species Specific ◽  
Inside Out ◽  
Positively Charged

The human apoptosis channel TRPM2 is stimulated by intracellular ADR-ribose and calcium. Recent studies show pronounced species-specific activation mechanisms. Our aim was to analyse the functional effect of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), commonly referred to as PIP2, on different TRPM2 orthologues. Moreover, we wished to identify the interaction site between TRPM2 and PIP2. We demonstrate a crucial role of PIP2, in the activation of TRPM2 orthologues of man, zebrafish, and sea anemone. Utilizing inside-out patch clamp recordings of HEK-293 cells transfected with TRPM2, differential effects of PIP2 that were dependent on the species variant became apparent. While depletion of PIP2 via polylysine uniformly caused complete inactivation of TRPM2, restoration of channel activity by artificial PIP2 differed widely. Human TRPM2 was the least sensitive species variant, making it the most susceptible one for regulation by changes in intramembranous PIP2 content. Furthermore, mutations of highly conserved positively charged amino acid residues in the membrane interfacial cavity reduced the PIP2 sensitivity in all three TRPM2 orthologues to varying degrees. We conclude that the membrane interfacial cavity acts as a uniform PIP2 binding site of TRPM2, facilitating channel activation in the presence of ADPR and Ca2+ in a species-specific manner.

Role of Intrahelical Arginine Residues in Functional Properties of Uncoupling Protein (UCP1)†

Biochemistry ◽  
2001 ◽  
Vol 40 (17) ◽  
pp. 5243-5248 ◽  
Author(s):  
Karim S. Echtay ◽  
Martin Bienengraeber ◽  
Martin Klingenberg
Keyword(s):  
Functional Properties ◽  
Uncoupling Protein ◽  
Arginine Residues

scholarly journals Characterization of the Drosophila protein arginine methyltransferases DART1 and DART4

2004 ◽  
Vol 379 (2) ◽  
pp. 283-289 ◽  
Author(s):  
Marie-Chloé BOULANGER ◽  
Tina Branscombe MIRANDA ◽  
Steven CLARKE ◽  
Marco di FRUSCIO ◽  
Beat SUTER ◽  
...  
Keyword(s):  
Rna Binding ◽  
Developmental Stages ◽  
Rna Binding Proteins ◽  
Genetic System ◽  
Arginine Methylation ◽  
Type I ◽  
Protein Arginine Methyltransferases ◽  
Arginine Residues ◽  
Drosophila Protein

The role of arginine methylation in Drosophila melanogaster is unknown. We identified a family of nine PRMTs (protein arginine methyltransferases) by sequence homology with mammalian arginine methyltransferases, which we have named DART1 to DART9 (Drosophilaarginine methyltransferases 1–9). In keeping with the mammalian PRMT nomenclature, DART1, DART4, DART5 and DART7 are the putative homologues of PRMT1, PRMT4, PRMT5 and PRMT7. Other DART family members have a closer resemblance to PRMT1, but do not have identifiable homologues. All nine genes are expressed in Drosophila at various developmental stages. DART1 and DART4 have arginine methyltransferase activity towards substrates, including histones and RNA-binding proteins. Amino acid analysis of the methylated arginine residues confirmed that both DART1 and DART4 catalyse the formation of asymmetrical dimethylated arginine residues and they are type I arginine methyltransferases. The presence of PRMTs in D. melanogaster suggest that flies are a suitable genetic system to study arginine methylation.

scholarly journals Role of glycerol 3-phosphate dehydrogenase in glyceride metabolism. Effect of diet on enzyme activities in chicken liver

1975 ◽  
Vol 146 (1) ◽  
pp. 223-229 ◽  
Author(s):  
J W Harding ◽  
E A Pyeritz ◽  
E S Copeland ◽  
H B White
Keyword(s):  
Lactate Dehydrogenase ◽  
High Fat Diet ◽  
Acyl Carrier Protein ◽  
Fatty Acid Synthetase ◽  
Carrier Protein ◽  
High Fat ◽  
Carbohydrate Diet ◽  
Metabolic Role ◽  
Glycerol 3 Phosphate Dehydrogenase

1. The metabolic role of hepatic NAD-linked glycerol 3-phosphate dehydrogenase (EC 1.1.1.8) was investigated vis-a-vis glyceride synthesis, glyceride degradation and the maintainence of the NAD redox state. 2. Five-week-old chickens were placed on five dietary regimes: a control group, a group on an increased-carbohydrate-lowered-fat diet, a group on a high-fat-lowered-carbohydrate diet, a starved group and a starved-refed group. In each group the specific activity (mumol/min per g wet wt. of tissue) of hepatic glycerol 3-phosphate dehydrogenase was compared with the activities of the β-oxoacyl-(acyl-carrier protein) reductase component of fatty acid synthetase, glycerol kinase (EC 2.7.1.30) and lactate dehydrogenase (EC 1.1.1.27). 3. During starvation, the activities of glycerol 3-phosphate dehydrogenase, glycerol kinase and lactate dehydrogenase rose significantly. After re-feeding these activities returned to near normal. All three activities rose slightly on the high-fat diet. Lactate dehydrogenase activity rose slightly, whereas those of the other two enzymes fell slightly on the increased-carbohydrate-lowered-fat diet. 4. The activity of the β-oxoacyl-(acyl-carrier protein) reductase component of fatty acid synthetase, a lipid-synthesizing enzyme, contrasted strikingly with the other three enzyme activities. Its activity was slightly elevated on the increased-carbohydrate diet and significantly diminished on the high-fat diet and during starvation. 5. The changes in activity of the chicken liver isoenzyme of glycerol 3-phosphate dehydrogenase in response to dietary stresses suggest that the enzyme has an important metabolic role other than or in addition to glyceride biosynthesis.

Role of active site arginine residues in substrate recognition by PPM1A

2021 ◽  
Author(s):  
Itsumi Tani ◽  
Shogo Ito ◽  
Yukiko Shirahata ◽  
Yutaka Matsuyama ◽  
James G. Omichinski ◽  
...  
Keyword(s):  
Active Site ◽  
Substrate Recognition ◽  
Arginine Residues

scholarly journals Effects of modifications of α-crystallin on its chaperone and other properties

2002 ◽  
Vol 364 (3) ◽  
pp. 711-717 ◽  
Author(s):  
Barry K. DERHAM ◽  
John J. HARDING
Keyword(s):  
Congenital Cataract ◽  
Point Mutations ◽  
Small Heat Shock Protein ◽  
High Molecular Mass ◽  
Lens Crystallins ◽  
Post Translational Modifications ◽  
Chaperone Function ◽  
Arginine Residues

The role of α-crystallin, a small heat-shock protein and chaperone, may explain how the lens stays transparent for so long. α-Crystallin prevents the aggregation of other lens crystallins and proteins that have become unfolded by ‘trapping’ the protein in a high-molecular-mass complex. However, during aging, the chaperone function of α-crystallin becomes compromised, allowing the formation of light-scattering aggregates that can proceed to form cataracts. Within the central part of the lens there is no turnover of damaged protein, and therefore post-translational modifications of α-crystallin accumulate that can reduce chaperone function; this is compounded in cataract lenses. Extensive in vitro glycation, carbamylation and oxidation all decrease chaperone ability. In the present study, we report the effect of the modifiers malondialdehyde, acetaldehyde and methylglyoxal, all of which are pertinent to cataract. Also modification by aspirin, which is known to delay cataract and other diseases, has been investigated. Recently, two point mutations of arginine residues were shown to cause congenital cataract. 1,2-Cyclohexanedione modifies arginine residues, and the extent of modification needed for a change in chaperone function was investigated. Only methylglyoxal and extensive modification by 1,2-cyclohexanedione caused a decrease in chaperone function. This highlights the robust nature of α-crystallin.

scholarly journals Role of the N-terminal region in covalent modification of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase: comparison of phosphorylation and ADP-ribosylation

1995 ◽  
Vol 309 (1) ◽  
pp. 119-125 ◽  
Author(s):  
J L Rosa ◽  
J X Pérez ◽  
F Ventura ◽  
A Tauler ◽  
J Gil ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Covalent Modification ◽  
Terminal Region ◽  
Bifunctional Enzyme ◽  
Dependent Protein Kinase ◽  
Adp Ribosylation ◽  
Camp Dependent Protein Kinase ◽  
Arginine Residues ◽  
Dependent Protein

The effect of cyclic AMP (cAMP)-dependent phosphorylation and ADP-ribosylation on the activities of the rat liver bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2/FBPase-2), was investigated in order to determine the role of the N-terminus in covalent modification of the enzyme. The bifunctional enzyme was demonstrated to be a substrate in vitro for arginine-specific ADP-ribosyltransferase: 2 mol of ADP-ribose was incorporated per mol of subunit. The Km values for NAD+ and PFK-2/FBPase-2 were 14 microM and 0.4 microM respectively. A synthetic peptide (Val-Leu-Gln-Arg-Arg-Arg-Gly-Ser-Ser-Ile-Pro-Gln) corresponding to the site phosphorylated by cAMP-dependent protein kinase was ADP-ribosylated on all three arginine residues. Analysis of ADP-ribosylation of analogue peptides containing only two arginine residues, with the third replaced by alanine, revealed that ADP-ribosylation occurred predominantly on the two most C-terminal arginine residues. Sequencing of the ADP-ribosylated native enzyme also demonstrated that the preferred sites were at Arg-29 and Arg-30, which are just N-terminal to Ser-32, whose phosphorylation is catalysed by cAMP-dependent protein kinase (PKA). ADP-ribosylation was independent of the phosphorylation state of the enzyme. Furthermore, ADP-ribosylation of the enzyme decreased its recognition by liver-specific anti-bifunctional-enzyme antibodies directed to its unique N-terminal region. ADP-ribosylation of PFK-2/FBPase-2 blocked its phosphorylation by PKA, and decreased its PFK-2 activity, but did not alter FBPase-2 activity. In contrast, cAMP-dependent phosphorylation inhibited the kinase and activated the bisphosphatase. These results demonstrate that ADP-ribosylation of arginine residues just N-terminal to the site phosphorylated by PKA modulate PFK-2 activity by an electrostatic and/or steric mechanism which does not involved uncoupling of N- and C-terminal interactions as seen with cAMP-dependent phosphorylation.

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