Tariq Mustafa

Metabolic sources of power for mantle muscle of a fast swimming squid

Abstract 1. 1. Squid mantle muscle utilizes carbohydrate as its primary carbon and energy source. 2. 2. Lactate does not accumulate under any circumstances because of a lack of lactate dehydrogenase. Instead, during imposed anoxic stress, alpha-glycerophosphate + pyruvate or metabolic derivatives of pyruvate accumulate in about a 1:1 ratio. 3. 3. The muscle is rich in mitochondria and displays high activities of citrate synthase, malate dehydrogenase, and aspartate aminotransferase. 4. 4. High α-glycerophosphate dehydrogenase activity in the cytoplasm and high rates of mitochondrial α-glycerophosphate oxidase indicate the the potential for an active α-glycerophosphate cycle comparable to that known in insect flight muscle. 5. 5. These characteristics, in addition to consistent control properties of several key regulatory enzymes, support the hypothesis of an obligatorily aerobic carbohydrate catabolism in squid mantle muscle.

Enzymes in facultative anaerobiosis of molluscs—III. Phosphoenolpyruvate carboxykinase and its role in aerobic-anaerobic transition

Abstract 1. 1. In this paper, the regulatory properties of p-enolpyruvate carboxykinase (E.C. 4.1.1.32) from oyster adductor tissue are further studied. 2. 2. Competitive inhibition patterns are obtained for metal ions with ITP as the inhibitor. In both instances 0·25 mM ITP increases the apparent K m values of the metal ion by tenfold. 3. 3. In the presence of Zn 2+ with ITP as the inhibitor and IDP as the variable substrate, a linear non-competitive inhibition pattern is obtained. At all concentrations of ITP tested the K m (IDP) remains unchanged at about 0·06 mM. In contrast in the presence of Mn 2+ a linear, mixed competitive ITP inhibition pattern is obtained. In this instance, 0·25 mM ITP increases the K m (IDP) several fold. 4. 4. In the presence of either metal ion, ITP inhibition is competitive with respect to p-enolpyruvate. With Zn 2+ , 0·5 mM ITP increases the t m (p-enolpyruvate) by about twofold while in the presence of Mn 2+ , 0·25 mM ITP increases the K m (p-enolpyruvate) at least by fivefold. 5. 5. In the presence of IDP or GDP, GTP also inhibits the p-enolpyruvate carboxykinase reaction. 6. 6. Alanine has a slight stimulatory effect at low p-enolpyruvate concentrations and markedly deinhibits ITP (or GTP) inhibited p-enolpyruvate carboxykinase. 7. 7. From the data presented it is speculated that co-ordinated changes in the intracellular concentrations of H + , ITP, GTP and alanine control p-enolpyruvate carboxykinase activity in vivo .

Enzymes in facultative anaerobiosis of molluscs. I. Malic enzyme of oyster adductor muscle.

1. 1. Malic enzyme (E.C. 1.1.1.40) in the adductor muscle of the Pacific oyster, Crossostrea gigas, occurs as a single electrophoretic species. 2. 2. Pyruvate saturation curves for the enzyme reaction are hyperbolic with an apparent Km for pyruvate of about 14–15 mM. Malate saturation curves are sigmoidal (Hill coefficient of 2·0) with S0·5 values of 0·4–0·5 mM, indicating about a thirtyfold greater affinity of malic enzyme for malate than for pyruvate. 3. 3. In the forward direction, malic enzyme is not product-inhibited by pyruvate, but in contrast, in the backward direction the enzyme is potently product-inhibited by malate. 4. 4. NADPH and NADH also strongly inhibit enzyme activity in the direction of malate decarboxylation and the inhibition is competitive with respect to NADP. In terms of steepness of the response and the maximum inhibition attained, NADH is a more efficient inhibitor of the enzyme at pH values above neutrality. Acidification in effect leads to a loss of this regulatory characteristic. 5. 5. From these studies it is concluded that in vivo malic enzyme functions mainly in the malate → pyruvate direction and that its activity is regulated by the pH and the redox potential of the cell. 6. 6. The metabolic functions of malic enzyme in the overall anaerobic metabolism of oyster muscle are briefly discussed.

Enzymes in facultative anaerobiosis of molluscs. II. Basic catalytic properties of phosphoenolpyruvate carboxykinase in oyster adductor muscle.

Abstract 1. 1. p-Enolpyruvate carboxykinase (E.C. 4.1.1.32) from oyster adductor tissue occurs as a single activity peak with a pI value of 6·64. 2. 2. The enzyme is present in high activities only in adductor tissue. 3. 3. It exhibits a relatively high degree of specificity for IDP and GDP and is inactive in the presence of IMP, GMP and ADP. Oyster enzyme, unlike p-enolpyruvate carboxykinases from other sources, exhibits no activity in the presence of Mg 2+ . 4. 4. The pH activity profiles in the presence of Zn 2+ and Mn 2+ are similar in general pattern but the pH optima are 5·1 and 6·0 respectively. 5. 5. Kinetic constants for metal ions, IDP and p-enolpyruvate were determined at different pH values. For metal ions, the apparent enzyme-Mn 2+ affinity is greater at pH 6·0 than at pH 5·1 while the apparent enzyme-Zn 2+ affinity is greater at pH 5·1 than at pH 6·0. S 0·5 values for both metal ions under different conditions vary between 0·15 and 0·52 mM. The absolute values of S 0·5 for IDP are comparable in the presence of Zn 2+ and Mn 2+ (in the range of 0·03–0.06 mM) and these are largely independent of pH. p-Enolpyruvate saturation exhibits aberrant behaviour in the present of Mn 2+ while in the presence of Zn 2+ it follows Michaelis-Menten patterns. Equally significant, the apparent K m values are at least one order of magnitude less than observed for the enzyme in the presence of Mn 2+ . 6. 6. Low concentrations of Cu 2+ (10–40 μM) inhibit the enzyme in the presence of either Zn 2+ or Mn 2+ . Cu 2+ in the presence of Mn 2+ exhibits a competitive inhibition pattern while in the presence of Zn 2+ , a non-competitive inhibition pattern occurs. 7. 7. The physiological role of the enzyme is discussed in relation to its cellular distribution, metal ion requirements and pH properties.

Squid muscle malic enzyme

Abstract 1. 1. Malic enzyme occurs in squid mantle muscle at low specific activity; it appears to function only in the forward direction, displaying a neutral pH optimum for pyruvate formation. 2. 2. Arrhenius plots between 5° and 30°C are linear and the reaction displays a temperature coefficient of about 4·0. 3. 3. Enzyme activity shows a pressure optimum at 100–200 atm at both low and high temperatures. 4. 4. Saturation kinetics follow Michaelis-Menten theory. 5. 5. In vivo the enzyme activity is probably integrated with the redox state of the cell, for malic enzyme catalysis is strongly inhibited by NADPH. 6. 6. NADPH inhibition is greatly potentiated by low temperature.

The lactacid oxygen debt in frogs after one hour's apnoea in air

Summary1.During one hours apnoea in air curarisedRana pipiens showed a reduction in oxygen metabolism of some 21 %. When artificial ventilation was restarted the frogs consumed 178 μl more oxygen in the first 10 min than in comparable periods before the apnoea. Over the recovery period of one hour the frogs consumed 350 μl more oxygen than in the hour before the apnoea.2.Blood lactic acid concentrations increased during apnoea and fell during the recovery period, the highest lactate levels being attained at the end of the apnoea and not during the initial part of the recovery period. The lactate: pyruvate ratio rose during apnoea. Blood glucose concentrations were above pre-apnoeic values in the early part of the recovery period.3.After one hours apnoea in oxygen, during which there was no significant reduction in oxygen metabolism, curarised frogs displayed no oxygen debt when artificial ventilation was restored.4.The results of the present experiments are discussed in the light of previous data and it is concluded that, in normal frogs, the increase in oxygen uptake following submergence has three elements, the major one being the oxygen cost of increased activity in the form of hyperventilation.