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Y S Brownstone

Publications and source records attributed to Y S Brownstone.

6 recordsLinked to original sources
PubMed1976

A study of phosphoglycerate kinase in human erythrocytes. I. Enzyme isolation, purification and assay.

The enzyme ATP-3-phospho-D-glycerate-1-phosphotransferase (EC 2.7.2.3) (phosphoglycerate kinase) has been isolated from human red cells in crystalline form by a modification of the method of Yoshida and Watanabe (1972) J. Biol. Chem. 247, 440-445). The crystalline enzyme was further purified by electrofocusing using carrier ampholytes (pH 7-9). The isoelectric point of phosphoglycerate kinase was estimated to be 8.75. The specific activity of purified phosphoglycerate kinase from electrofocusing was 2200 units per mg of protein at pH 8.3 (37 degrees C). Enzyme activity was assayed in the forward direction leading from 1,3-diphosphoglycerate to a 3-phosphoglycerate using a fluorimetric procedure for NAD-coupled enzymes for the measurement of the reaction rate at very low substrate concentrations. The auxiliary indicator enzymes were added in excess to yield true initial velocity kinetics, i.e. with no time lag upon addition of substrate (1,3-diphosphoglycerate). This was established theoretically using a mathematical model and confirmed experimentally. Further phosphoglycerate kinase was shown to be the rate-limiting step when the assay conditions were varied.

Adenosine Diphosphate↗
PubMed1976

A study of phosphoglycerate kinase in human erythrocytes. II. Kinetic properties.

Kinetic studies on phosphoglycerate kinase (EC 2.7.2.3) were performed in the forward reaction leading from 1,3-diphosphoglycerate to 3-phosphoglycerate. Substrate activation was observed at fixed levels of ADP or Mg2+ and varying concentrations of 1,3-diphosphoglycerate. A biphasic curve was obtained in both linear and double reciprocal plots demonstrating two Km values (Km1 1.9 - 10(-6) and Km2 9.8 - 10(6) M). Michaelis-Menten-type kinetics were observed in both the linear and double reciprocal plots at fixed levels of 1,3-diphosphoglycerate of ADP and varying concentrations of Mg2+. Apparent Michaelis-Menten kinetics were observed in linear plots when conditions of fixed concentrations of 1,3-diphosphoglycerate or Mg2+ were maintained with varying concentrations of ADP. However, the double-reciprocal plots demonstrated biphasic curves with two Km values (Km1 1.7 - 10(-5) and Km2 1.0 - 10(-4)M). Apparent negative cooperativity was observed with respect to 1,3-diphosphoglycerate and ADP. Phosphoglycerate kinase activity was found to be inhibited by AMP and 2,3-diphosphoglycerate. Substrate activation by 1,3-diphosphoglycerate was maintained in the presence of AMP or 2,3-diphosphoglycerate but at a reduced level of enzyme activity. AMP was found to inhibit enzyme activity non-competitively with respect to 1,3-diphosphoglycerate, ADP and Mg2+. 2,3-Diphosphoglycerate inhibits phosphoglycerate kinase activity with respect to 1,3-diphosphoglycerate, ADP and Mg2+. 2,3-Diphosphoglycerate inhibits phosphoglycerate kinase activity non-competitively with respect of 1,3-diphosphoglycerate.

Adenosine Diphosphate↗
PubMed1976

The reduction of coma time in lipophilic drug overdose using castor oil.

A clinical trial of castor oil in overdoses of lipophilic drugs gave a strong clinical impression that it was effective in speeding up recovery. Therefore, animal experiments were undertaken to confirm that castor oil acts as a ligand in Ethchlorvynol poisoning and that its use reduces coma time. Serial serum levels of Ethchlorvynol were obtained from dogs given Ethchlorvynol 150 mg/kg alone, and the same dose dissolved in castor oil 15 ml/kg in a crossover fashion. The result was a reduction of peak serum levels and of the half-life of the drug when the castor oil solution was used. In order to mimic the clinical situation more closely, a further crossover study was undertaken using Ethchlorvynol 300 mg/kg alone and the same dose followed by castor oil 15 ml/kg repeated q12h. This showed no delay in reaching peak serum concentration and no reduction of peak levels. However, it did show a 31 per cent reduction in the half-life of the drug. This change is statistically significant, and supports the continued use of castor oil in lipophilic drug overdose.

Animals↗