Enzyme dephosphorylation

Baddiley and coworkers42 have studied the structure of S13, which is composed of D-galactose, D-glucose, 2-acetamido-2-deoxy-D-glucose, and ribitol residues, and phosphate groups in the molar proportions 2 1 1 1 1. O-Acetyl groups are also present. A neutral pentasaccharide was obtained by hydrolysis with alkali, followed by enzymic dephosphorylation. On mild, acid hydrolysis, this yielded two main products, a trisaccharide (19) and a disaccharide (20), the structures of which were determined by conventional methods. 

Hydroxy-3-methylglutaryl (HMG)-CoA reductase on the smooth endoplasmic reticulum (SER) is the rate-limiting enzyme. Insulin acth"ates the enzyme (dephosphorylation), and glucagon inhibits it. Mevalonate is the product, and the statin drugs competitively inhibit the enzyme. Cholesterol represses the expression o the HMG-CoA reductase gene and also increases degradation of the enzyme. 

Two key enzymes in the pathway are regnlated by interconversion cycles they are the regnlatory enzyme PFK-2, and the glycolytic enzyme pyruvate kinase. There are two separate protein kinases that phosphorylate these enzymes and they both resnlt in activation of these enzymes. Dephosphorylation inactivates them. 

The exterior of the myocardial enzyme, situated in the plasmalemma, is considered to be the specific binding site of cardiac steroid glycosides. It is believed that the positive inotropic effect is due to the inhibition of enzyme dephosphorylation and... 

A small proportion of O-D-glucosylribitol was produced directly by hydrolysis of the teichoic acid with alkali ( see Fig. 16) this product is identical with that obtained by dephosphorylation of the hydrolysis mixture. The major products of such a hydrolysis with alkali were the isomeric monophosphates (58) and (59), in which R = 0-D-glucopyranosyl, both of which gave the O-D-glucosylribitol on enzymic dephosphorylation. The isomer (58) reduced 3 molar proportions of periodate, and the ribitol residue was oxidized, whereas the isomer (59) reduced 2 molar proportions of periodate, the ribitol residue being resistant to oxidation. Small proportions of the diphosphates (56) and (57) were also produced. Oxidation of the diphosphate (57) with periodate, followed by treatment with alkali to remove the aldehydic residues, gave a ribitol diphosphate. 

How is the enzyme dephosphorylated and activated Insulin stimulates the carboxylase by causing its dephospkorylation. It is not clear which of the phosphatases activates the carboxylase in response to insulin. The hormonal control of acetyl CoA carboxylase is reminiscent of that of glycogen synthase (Section 21.5.2). 

A similar result is obtained by heating the phospholipid in diphenyl ether with a trace of water [285] or by enzymic dephosphorylation [286]. The diglycerides produced can be silylated and both the location on the glycerol chain and the identity of the fatty acids determined by GC-MS. 

Gulland and Jackson performed some experiments with 5-nucleotidase, a highly specific enzyme which dephosphorylates 5-phospho-adenosine and -inosine but not" 5-phospho-guanosine and -uridine it is apparently not yet known whether the enzyme dephosphorylates 5-phos-pho-cytidine. They found that a mixture of phosphodiesterase with 5-nucleotidase liberates 35% of the total phosphorus as inorganic phosphate, and therefore decided that two or more of the phosphoryl groups may be attached at position (5) of the ribose units. The 35% dephosphorylation, intermediate between 25 and 50%, was explained as the result of simultaneous, competitive diesterase action at A and B, on two or more phosphoryl groups ... 

Enzyme activity can be regulated by covalent modification or by noncovalent (allosteric) modification. A few enzymes can undergo both forms of modification (e.g., glycogen phosphorylase and glutamine synthetase). Some covalent chemical modifications are phosphorylation and dephosphorylation, acetylation and deacetylation, adeny-lylation and deadenylylation, uridylylation and deuridyly-lation, and methylation and demethylation. In mammalian systems, phosphorylation and dephosphorylation are most commonly used as means of metabolic control. Phosphorylation is catalyzed by protein kinases and occurs at specific seryl (or threonyl) residues and occasionally at tyrosyl residues these amino acid residues are not usually part of the catalytic site of the enzyme. Dephosphorylation is accomplished by phosphoprotein phosphatases ... 

The non-enzymic dephosphorylation of O-phosphorothreonine which is brought about by pyridoxal in aqueous media has been investigated and a mechanism for the reaction has been proposed (Scheme 2). Copper(ii) and oxovanadium(iv) ions exert a strong catalytic effect and the dephosphorylation proceeds with C—O fission. The initial formation of a Schiff base may occur, followed by the loss of a proton from the -carbon atom of the threonine. 0-Phosphoro-a-methylserine, which does not possess an a-proton, does not dephosphorylate readily in aqueous solution. 

Phosphoenolpyruvate.—Phosphoenolpyruvate carboxytransphosphorylase catalyses two separate conversions of phosphoenolpyruvate (6) (Scheme 3). In the absence of carbon dioxide pyruvate and inorganic pyrophosphate may be formed enzymic dephosphorylation of (6) to enolpyruvate is probably followed by the non-enzymic protonation of the latter giving rise to pyruvate. 

Carbohydrate SXA, on acidic hydrolysis, yields galactose, galactose phosphate, galactosamine, ribitol, anhydroribitol, ribitol phosphates, and inorganic phosphate. Alkaline hydrolysis of carbohydrate SXA, followed by enzymic dephosphorylation and separation of the product... 

Treatment of SXIA, after the consumption of one mole of periodate per mole, with phenylhydrazine, followed by acidic hydrolysis, yields a sugar phosphate which, on enzymic dephosphorylation, gives glucose. A galacto-sylerythritol is obtained from a Smith degradation of reduced, alkali-degraded SXIA. 

Finally, synthesis of oligonucleotides by degradation Homologues of oligouridylate have been prepared by successive digestion of RNA with RNase Ti and RNase U2, followed by deamination, to afford oligouridylate sequences with 3 -terminal purine nucleotides, which were subjected to enzymic dephosphorylation, periodate oxidation and P-elimination of the terminal residue, and a second dephosphorylation step.325 xhis afforded (Up)nU species (n = 1 - 7), which were separated chromatographically. 

An additional example of how enzymes can affect the physical properties of purified proteins relates to the enzymic dephosphorylation of /3-casein (4). Bovine /3-casein contains five phosphate groups per monomer as phosphoseryl residues. Purified /3-casein from bovine milk was dephosphorylated by a phosphoprotein phosphatase. Both the native /3-casein and 65% dephosphorylated /3-casein self-associated to form polymers at 35°C. However, the dephosphorylated /3-casein had a larger sedimentation coefficient (S35 22.5) than that of the native /3-casein (S35 18.2). Also the sedimentation pattern of the dephosphorylated /3-casein was more polydisperse than the hypersharp pattern of the native /3-casein. These properties were accentuated with 95% dephosphorylated /3-casein. The decrease in the negative charge caused by the loss of phosphate apparently favors the self-association of /3-casein resulting from hydrophobic bonding. 

Horita, A. Weber, L.J. 1961. The enzymic dephosphorylation and oxidation of psilocybin and psilocin hy mammalian tissue homogenates. Biochemical Pharmacology Vol. 7 47-54. 

Protein Serine/Threonine Phosphatases. As their name implies, these enzymes dephosphorylate phosphoserine and phosphothreonine residues in proteins. They are metallophosphatases, with binuclear metal centers at their active sites, in common with the PAPs. Based on their catalytic domains they are grouped into two subfamilies, designated as PPP and PPM (40,41). 

GC, generally chromatography with a carrier gas (hydrogen, helium or nitrogen) as the mobile phase. Useful for any volatile lipid compound, such as fatty acid methyl esters, triacylglycerols and sterol esters, or compounds that can be made volatile, such as phospholipids after (enzymic) dephosphorylation. 

---