Synthesis of Peptides in Enzymic Reactions involving Glutathione

y-Glutamyl transpeptidase was isolated from sheep kidney cortex as an apparently homogeneous, highly active protein. At optimal pH and in the absence of acceptors, the enzyme catalyzes the release of about 510 pmol of p-nitroaniline per mg protein per min from the model substrate L-y-glutamylp-nitroanilide. Polyacrylamide gel electrophoresis in a sodium dodecylsulfate buffer system showed the presence of a large ( M , z 65000) and a small ( M , = 27000) polypeptide chain. Dissociation into two polypeptide chains was also achieved in 8 M urea. Amidination with dimethylsuberimidate produced a crosslinked protein of molecular weight approximately 90000.In the course of this work a convenient procedure was developed for the determination of y-glutamyl transpeptidase activity using ~-[gfycine-2-~H]glutathione as the substrate. In this procedure the release of ~ysteinyl-[2-~H]glycine from glutathione is followed, after separation of the radioactive dipeptide from unreacted glutathione on a small Dowex-1 acetate column. The reactions with y-glutamyl-p-nitroanilide and glutathione are both strongly activated by several metal ions (Ca2 +, Mg2 + , Na+ and K + ) and by a number of amino acids and peptide acceptors. The products of the reaction with glutathione were identified as cysteinylglycine, y-glutamylglutathione and glutamate. The formation of these products is consistent with the function of y-glutamyl transpeptidase in both the y-glutamyl transfer reaction and in the hydrolysis of the y-glutamyl bond. The activating effect of metal ions in the reaction with glutathione was shown to be dependent on the acceleration of the transfer reaction; the rate of hydrolysis of the y-glutamyl bond remaining unchanged.

show abstract

“…The enzyme is also capable of hydrolysing the y-glutamyl bond of gluta- (1) En:yme. 9-Glutamyl transpeptidaae (EC 2.3.2.2).…”

mentioning

confidence: 99%

gamma-Glutamyl Transpeptidase of Sheep-Kidney Cortex. Isolation, Catalytic Properties and Dissociation into Two Polypeptide Chains

Zelazo

Orłowski

1976

Eur J Biochem

View full text Add to dashboard Cite

show abstract

“…The enzyme is widely distributed in animal tissues, kidney being most active. It is notable that the enzyme is bound to tissue particles and that purified preparations of the enzyme contain as much as 30%0 carbohydrate.8 '9 -y-Glutamyl cyclotransferase (y-glutamyl lactamase), which catalyzes reaction (2)10,…”

mentioning

confidence: 99%

The γ-Glutamyl Cycle: A Possible Transport System for Amino Acids

Orłowski

Meister

1970

Proc. Natl. Acad. Sci. U.S.A.

473

218

View full text Add to dashboard Cite

Abstract. Evidence is presented that rat kidney contains enzymes that catalyze the synthesis and utilization of glutathione; these reactions, which involve the uptake and release of amino acids from y-glutamyl linkage, constitute a cyclical process which is termed "the y-glutamyl cycle." The y-glutamyl cycle has properties that fulfill the requirements of an amino acid transport system. Thus, -y-glutamyl transpeptidase may function in translocation and y-glutamylcysteine synthetase and glutathione synthetase may catalyze energy-requiring "recovery" steps in transport. These and other considerations suggest that glutathione serves a carrier function in amino acid transport.It has long been known that kidney preparations catalyze the degradation of glutathioneI, and that y-glutamyl transpeptidase2 plays an important role in this process. This enzyme catalyzes the transfer of the -y-glutamyl moiety of glutathione to a variety of a-amino acids:glutathione + amino acid --y-glutamylamino acid + cysteinylglycine (1)Purified preparations of -y-glutamyl transpeptidase also catalyze the liberation of glutamate from glutathione and -y-glutamylamino acids. The broad amino acid specificity of reaction (1) has been amply demonstrated3-7. The enzyme is widely distributed in animal tissues, kidney being most active. It is notable that the enzyme is bound to tissue particles and that purified preparations of the enzyme contain as much as 30%0 carbohydrate.8'9 -y-Glutamyl cyclotransferase (y-glutamyl lactamase), which catalyzes reaction (2)10,-y-glutamylamino acid --pyrrolidone carboxylate + amino acid (2) acts rapidly on the -y-glutamyl derivatives of glutamine, alanine, and glycine. When both transpeptidase (TP) and cyclotransferase (CT) are present the y-glutamyl derivatives of virtually all of the protein amino acids are degraded by the following pathway11: 'y-Glu-amino acid + y-Glu-amino acid TP y-Glu-y-Glu-amino acid + amino acid (3a) 7y-Glu-y-Glu-amino acid CT pyrrolidone carboxylate + y-Glu-amino acid (3b) 1248

show abstract

“…y-Glutamyltranspeptidase activity has been demonstrated in animal tissues (Hanes, Hird & Isherwood, 1950) and in fungi (Waelisch, 1952).…”

Section: Discussionmentioning

confidence: 99%

Sulphur Metabolism of Puccinia graminis f.sp. tritici in Axenic Culture

Howes

Scott

1973

Journal of General Microbiology

View full text Add to dashboard Cite

SUMMARYAxenic cultures of Puccinia graminis f. sp. tritici cannot reduce sulphate for sulphur-amino-acid synthesis. Although [35S]sulphide is incorporated into these amino acids, the fungus still has a nutritional requirement for a sulphur amino acid. Most of the incorporated sulphide was found in the culture filtrate in the form of cysteine, S-methylcysteine, glutathione and cysteinylglycine. There were appreciable amounts of labelled methionine in mycelial protein but very little of this amino acid appeared in the culture filtrate. The results indicate that the nutritional requirements for sulphur amino acids by the rust fungus is caused by loss of cysteine and some of its derivatives into the medium.

show abstract

Synthesis of Peptides in Enzymic Reactions involving Glutathione

Cited by 308 publications

References 19 publications

gamma-Glutamyl Transpeptidase of Sheep-Kidney Cortex. Isolation, Catalytic Properties and Dissociation into Two Polypeptide Chains

gamma-Glutamyl Transpeptidase of Sheep-Kidney Cortex. Isolation, Catalytic Properties and Dissociation into Two Polypeptide Chains

The γ-Glutamyl Cycle: A Possible Transport System for Amino Acids

Sulphur Metabolism of Puccinia graminis f.sp. tritici in Axenic Culture

Contact Info

Product

Resources

About