Separation and Purification of Enzymes Associated with Insoluble Particles

Morton, R. K.

doi:10.1038/1661092a0

Cited by 268 publications

(55 citation statements)

References 23 publications

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“…Cohn Fractions II,3 III and IV-1 were subjected to lipid extraction by several methods: (A) ethanol-ether (3:1) in a one-phase system at 600 C. for three 10 minute periods, (B) n-butanol in a two-phase system with water at room temperature overnight (11), (C) n-butanol in a one-phase system at -150 C. (11). In each instance the lipid solvent layer was removed to tared vessels for drying and weighing of lipid residues, the residue was extracted in water which was then removed and subjected to rotary evaporation at 460 C. and measured for hexuronic acid content, and the residue itself was subjected to enzymatic hydrolysis and precipitation with cetyl trimethyl ammonium bromide and measurement of hexuronic acid content.…”

Section: Methodsmentioning

confidence: 99%

The Distribution of Acid Mucopolysaccharides in Cohn Fractions of Human Plasma Proteins1

Kerby¹

1958

J. Clin. Invest.

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Section: Methodsmentioning

confidence: 99%

The Distribution of Acid Mucopolysaccharides in Cohn Fractions of Human Plasma Proteins1

Kerby¹

1958

J. Clin. Invest.

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“…I t catalyzes both the hydrolysis of y glutamyl-containing peptide bonds and the transfer of y glutamyl groups from donor peptides to suitable acceptors, i. e. transpeptidation. GGTP activity has been found in many tissues [3, 1 1, 13,23,27] of small laboratory animals [ll], hog [26], cow [20], sea lion [2], and man 13,11,13,23,271. Techniques originally developed for estimating GGTP activity involved quantitating the disappearance of glutathione and the appearance of the products of glutathione cleavage [4,8,28].…”

Section: Introductionmentioning

confidence: 99%

Gamma Glutamyl Transpeptidase: Measurement and Development in Guinea Pig Small Intestine

et al. 1969

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ExtractA modification ofa previous method for the assay ofy glutamyI transpeptidase (GGTP) was developed.Substrate solubility difficulties alluded to by other investigators were avoided by employing heating and solubilization of the chromogenic substrate y glutamyl-P-naphthylamide in a medium of carbonate 0.05 M and Tris buffer 0.1 M a t p H 9.5. The kinetics and conditions for such an assay are described. Whole intestinal homogenates of adult male guinea pigs were used as the source of the enzyme.The developmental pattern of this enzyme was determined in fetuses at 55 and 63 days of gestation and at varying times from 1 to 90 days of age. A total of 69 animals was assayed. The general pattern was that ofhigh specific activity during prenatal life, with a rapid decline during the neonatal period (3-24 days of age) and a slight increase after 55 days of age.Other guinea pig organs were studied. Liver and kidney were found to contain enzyme activity greater than that of the intestine. Subcellular fractionation of intestinal mucosa using ultracentrifugation revealed a twenty-fold enrichment of activity in jejunal brush border membrane, compared with whole jejunal homogenates when expressed as specific enzyme activity per mg of protein. The stability characteristics of GGTP disclosed no loss of specific activity when stored a t -28' for 50 days.This simple enzyme assay, stability of the enzyme when frozen, subcellular distribution in the intestinal brush border membrane, and an unusual developmental pattern made this enzyme a useful adjunct to the study of intestinal protein metabolism. SpeculationThe unique ability of GGTP to hydrolyze y glutamic acid-peptide bonds and the location of the enzyme in the intestinal brush border suggest a role for this enzyme in the metabolism of an unusual group of physiologically important peptides such as glutathione, folic acid, and gluten-gliadin. The significance of these y-bonded compounds may now be approached in order to investigate the role of this enzyme in gluten enteropathy and related disorders.

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“…The rapid and complete inactivation of virus by n-butanol may be related to its reported efficiency in extracting phospholipids (Morton, 1950). The actions of diethyl ether and deoxycholate were studied separately by Andrewes & Horstmann (1949) and Burnet & Lush (1940), and the results were compared by Sunaga et al (1960), who added several findings and concluded that the two agents were strikingly similar in their actions on viruses.…”

Section: Discussionmentioning

confidence: 99%

The Action of Phospholipase A and Lipid Solvents on Murray Valley Encephalitis Virus

Anderson

Ada

1961

Journal of General Microbiology

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SUMMARYPhospholipase A (100 ,ug./ml.) acting at 37" for 30 min. decreased the infective titre of crude or purified Murray Valley encephalitis ( W E ) virus from 7 or 8 log to less than 1 log. This inactivated virus yielded 2 log of infective 'ribonucleic acid' ('RNA') when it was treated with phenol. Ether, chloroform or butanol acting at 0-4' each decreased the infectivity of crude or purified MVE virus. Again the inactivated virus yielded 'RNA' with an infective titre higher than the titre of the inactivated virus. Infective 'RNA' prepared from crude W E virus was not inactivated by ether, chloroform or butanol. High concentrations of phospholipase A preparations destroyed ' RNA', possibly due to traces of contaminating ribonuclease. The results suggest that intact viral phospholipid is necessary for infectivity of the virus particle.

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Separation and Purification of Enzymes Associated with Insoluble Particles

Cited by 268 publications

References 23 publications

The Distribution of Acid Mucopolysaccharides in Cohn Fractions of Human Plasma Proteins1

The Distribution of Acid Mucopolysaccharides in Cohn Fractions of Human Plasma Proteins1

Gamma Glutamyl Transpeptidase: Measurement and Development in Guinea Pig Small Intestine

The Action of Phospholipase A and Lipid Solvents on Murray Valley Encephalitis Virus

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