390. The standardisation of some buffer solutions at elevated temperatures

Britton, H. T. S.; Welford, George A.

doi:10.1039/jr9370001848

Cited by 50 publications

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“…Although the emf measurements of the solutions were made at room temperature, while the solubility experiments were made in most cases at higher temperatuJ'es, no account was taken of the small shift in pH of the buffer which accompanied this temperature change [6,7]. Since all electrode measurements were made in Pyrex beakers, the solubility tests were also carried out in similar vessels, even though Pyrex is known to be attacked by the alkaline solutions [8,9].…”

Section: Methodsmentioning

confidence: 99%

Effect of the solubility of glass on the behavior of the glass electrode

Hubbard¹,

Hamilton²,

Finn³

1939

J. RES. NATL. BUR. STAN.

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Section: Methodsmentioning

confidence: 99%

Effect of the solubility of glass on the behavior of the glass electrode

Hubbard¹,

Hamilton²,

Finn³

1939

J. RES. NATL. BUR. STAN.

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“…In both a universal barbitone buffer [35] and 50 mM Tris buffer the Nocardia enzyme displays a broad activity curve with a pH optimum of 8.4. Though during enzyme purification assays of activity Table5.…”

Section: Relationship Between P H and Enzyme Activitymentioning

confidence: 99%

The Purification and Properties of Cyclohexanone Oxygenase from Nocardia globerula CL1 and Acinetobacter NCIB 9871

Donoghue

Norris

Trudgill

1976

European Journal of Biochemistry

341

219

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Cyclohexanone oxygenases from Nocardia globerula CL1 and Acinetobacter NCIB 9571 have been purified 12‐fold and 35‐fold respectively and each gives a single symmetrical sedimentation peak in the ultracentrifuge and a single protein band on 2.25 nm average pore radius polyacrylamide gels. The enzyme from N. globerula has a molecular weight of 53000 while that from Acinetobacter has a molecular weight of about 59000. Each is a single polypeptide chain with one mole of bound FAD per mole of protein that does not dissociate during purification. Acidification of the Acinetobacter enzyme in the presence of (NH4)2SO4 releases the bound FAD and yields native apoenzyme from which the active holoenzyme can be reconstituted. The apparent dissociation constant for the FAD is 40 nM. The near unitary stoichiometry of cyclohexanone, NADPH and oxygen consumption is typical of mixed function oxygenases with external electron donors. The oxygenated product has been identified as 1‐oxa‐2‐oxocycloheptane thus placing these enzymes in the small group of lactone and ester‐forming oxygenases. Their correct systematic name is cyclohexanone. NADPH: oxygen oxidoreductase (1,2‐lactonizing) (EC 1.14.13.‐). A functionally essential sulfhydryl group is present at the catalytic centre of both enzymes but there is no reliable indication from inhibitor studies that they contain any functional metal ion. The three titratable sulfhydryl groups of the Acinetobacter enzyme are not equivalent since reaction with one of them selectively inhibits catalytic activity. Protection against sulfhydryl active agents is afforded by NADPH but not by cyclohexanone. The N. globerula enzyme has a pH optimum of 8.4, apparent Km values of 1.56 μM and 31.3 μM for cyclohexanone and NADPH respectively and a catalytic centre activity of 1018 ml substrate transformed × mol enzyme−1× min−1. The Acinetobacter enzyme has a pH optimum of 9.0, apparent Km values of 6.9 tM and 17.8 μM and a catalytic centre activity of 1390 mol × mol enzyme−1× min−1. Both enzymes display absolute specificity for electron donor which contrasts with the broad specificity for ketone substrate. An enzyme‐cyclohexanone complex has been detected by difference spectroscopy only in the case of the Nocardia enzyme. Rapid reduction of the enzyme‐bound FAD occurs upon addition of NADPH in the absence of cyclohexanone. Titration of enzyme with NADPH under anaerobic conditions and anaerobic photoreduction in the presence of EDTA have not revealed the formation of any stable flavin semiquinones. These enzymes bear a strong resemblance to several of the monooxygenases that hydroxylate aromatic compounds.

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“…3) was ob tained showing optima at about pH 6.5 and 9. These two pH optima were also observed when the experiment was repeated with one universal buffer system (25) in exchange for the different buffer systems indicated in fig ure 3.…”

Section: Resultsmentioning

confidence: 60%

Enzymatic Sulphation of Bile Salts in Man

Lööf¹

1981

Digestion

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Enzymatic sulphation of bile salts was identified in cytosol prepared from human adrenal tissue. Unconjugated lithocholate was sulphated at a slightly greater rate than glyco-lithocholate. Chenodeoxycholate and deoxycholate were sulphated at the 7- and 12-OH positions, respectively, but no disulphate formation from these bile salts was found. The physiological implications of these results are discussed.

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390. The standardisation of some buffer solutions at elevated temperatures

Cited by 50 publications

References 0 publications

Effect of the solubility of glass on the behavior of the glass electrode

Effect of the solubility of glass on the behavior of the glass electrode

The Purification and Properties of Cyclohexanone Oxygenase from Nocardia globerula CL1 and Acinetobacter NCIB 9871

Enzymatic Sulphation of Bile Salts in Man

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