Arthur R. Schulz

Thyroidal biosynthesis of iodothyronines: II. General characteristics and purification of mitochondrial monoamine oxidase☆

Abstract 1. 1. Monoamine oxidase (monoamine:O2 oxidoreductase, EC 1.4.3.4) has been partially purified from bovine thyroid mitochondria. The enzyme appeared to require Cu2+ and sulfhydryl groups for activity. Unlike the plasma oxidase, the thyroid enzyme was not inhibited by carbonyl reagents which suggests that the thyroid enzyme may not contain pyridoxal phosphate. Flavin analogs were inhibitory. 2. 2. Thyroid monoamine oxidase exhibited a rather high degree of substrate specificity. Of the amines tested, only tyramine and phenylethylamine were oxidized at an appreciable rate. 3-Iodotyramine was not oxidized, but was a potent non-competitive inhibitor. Enzymic activity was insensitive to 0.5 M urea, but this concentration of urea obviated iodotyramine inhibition. 3. 3. A tentative reaction mechanism has been proposed based on initial velocity and product inhibition studies.

Isolation and purification of rabbit adrenal norephinephrine N-methyl transferase isozymes

Abstract Five charge isozymes of rabbit adrenal norepinephrine N-methyl transferase have been isolated and purified to apparent homogeneity. The isolation and purification procedures include ammonium sulfate precipitation, diethylaminoethyl-cellulose chromatography, isoelectric focusing, and hydroxylapatite chromatography. Homogeneity was judged by disc gel electrophoresis. The isozymes appear to be monomeric charge isozymes with molecular weights in the range of 35,000 to 40,000. The ratio of activities of the five isozymes is different when isolated from the adrenal glands of young rabbits than when isolated from those of adult rabbits.

Product inhibition studies and the reaction sequence of rabbit adrenal norepinephrine N-methyl transferase isozymes.

Abstract The effects of reaction products on the steady-state kinetic properties of the five charge isozymes of rabbit adrenal norepinephrine N -methyl transferase have been investigated. Qualitative and quantitative differences were observed for the isozymes. The only characteristic that was common to all isozymes was the competition between S -adenosylmethionine and S -adenosylhomocysteine for the binding site. In most instances, the product inhibition constants were sufficiently low to suggest that product inhibition may be an important factor in regulating the activities of the isozymes. A reaction model is proposed for rabbit adrenal norepinephrine N -methyl transferase which is consistent with results observed in investigations of the steady-state kinetic properties of the five charge isozymes. The proposed model is that of an ordered sequential reaction sequence in which the active center contains a binding site for S -adenosylmethionine and S -adenosylhomocysteine, and a binding site for norepinephrine and epinephrine. The proposed model includes the formation of a number of abortive complexes between enzyme and substrate and product, but not all of the abortive complexes are significant kinetically in the case of some of the isozymes. The differences in the steady-state kinetic characteristics of the isozymes are attributed to differences in the magnitudes of the rate constants of some of the individual steps.

The interaction of rabbit adrenal norepinephrine N-methyl transferase isozymes with substrates

Abstract The steady-state kinetic behaviors of the five rabbit adrenal norepinephrine N -methyl transferase isozymes have been compared with particular reference to substrate inhibition patterns. Four distinct substrate inhibition patterns were observed. The E-1 isozyme was not subject to inhibition by either substrate, while the E-2 isozyme was inhibited by both substrates. The E-3 and E-4 isozymes were inhibited by norepinephrine only, while E-5 is inhibited only by S -adenosylmethionine. The substrate inhibition constants were sufficiently small in relation to the Michaelis constants to make substrate inhibition an important factor in regulation of activities of the isozymes.

Energy metabolism in the whole animal revisited

Dimensional analysis of animals with regard to those structures and functions which affect the rate of energy metabolism can be conducted on the basis of either geometric or elastic similarity. Predictions of the relationships between various structures and functions and body mass are dependent of the type of similarity assumed. Comparison of the relationships between numerous physiological variables and body mass reported in the literature and those predicted from geometric and elastic similarity indicates that the preponderance of predictions from elastic similarity are in better agreement with the observed values than are the predictions from geometric similarity. It appears that the components of the cardiovascular system are scaled in accordance with elastic similarity. For this reason, cardiac output is proportional to body mass raised to the 3/4 power. The biochemical basis for the relationship between cardiac output and the rate of energy metabolism is discussed.

Bovine thyroid cytosol glutamate dehydrogenase

Abstract The distribution of glutamate dehydrogenase in bovine thyroid tissue differs from the distribution observed in other mammalian tissues. While the mitochondria of thyroid tissue contain glutamate dehydrogenase which is ‘activated’ by treatment of the mitochondria with a non-ionic detergent, the majority of the glutamate dehydrogenase activity is present in the cytosol fraction. In the liver, both glutamate dehydrogenase and NAD-specific isocitrate dehydrogenase are found exclusively in the mitochondrial matrix. In contrast to thyroidal glutamate dehydrogenase, the NAD-specific isocitrate dehydrogenase was found exclusively in the mitochondrial fraction. Evidence is presented which indicates that the presence of glutamate dehydrogenase in thyroid cytosol is not due to rupture of the mitochondria. The possible physiological significance of thyroid cytosol glutamate dehydrogenase is discussed.

Nutrient-response: a long random walk through metabolic pathways

Abstract An equation has been derived to describe the relationship between response and nutrient intake. This derivation has been based on stochastic principles in which the metabolites involved were treated as discrete states. Derivation involved calculation of the fractional occupancy of each state during a continuous random walk through the graph, which represented the metabolic pathway. The resultant equation is a rational polynomial. A rational polynomial can accommodate the various shapes of nutrient-response curves that have been reported.

Chapter 15 – Nutrient–Response: A “Top Down” Approach to Metabolic Control

The chapter discusses a “top down” approach to metabolic control with reference to nutrient- responses. Nutrient-response curves contain information concerning the metabolic fate of the nutrient involved. The chapter describes an alternative that is the mathematical analysis of the nutrient-response curve in terms of the metabolic fate of the nutrient, and the integration of the analysis with the established principles developed in metabolic control theory and biochemical systems theory. It is important to recognize the essential relationship between the concepts of sensitivity and identifiability in mathematical modeling in nutrition. If the sensitivity is not readily identifiable, conditions must be sought to render the sensitivity identifiable. However, if the sensitivity is not identifiable readily, it is incumbent on the investigator to recognize the possible effect of such sensitivity. The metabolic pools defined in the denominator of the nutrient-response equation can be viewed as intermediate metabolites. Thus, the logarithmic derivative of each metabolic pool with respect to nutrient intake provides an estimate of the sensitivity of the pool to nutrient intake.

Effects of product inhibition in metabolic pathways: stability and control

The effect of product inhibition in metabolic pathways is examined using (a) an unbranched pathway in the absence of endproduct inhibition and (b) an unbranched pathway with endproduct inhibition. It is shown that product inhibition may be considered an alternative mechanism to endproduct inhibition for reducing the overall logarithmic gain of an unregulated pathway. When product inhibition and endproduct inhibition are both present, they act in concert with each other to lower the overall logarithmic gain and alleviate parameter sensitivities. Product inhibition is also found to exert a stabilizing influence that competes with the destabilizing effect of endproduct inhibition in controlling the dynamic behavior.

Kinetic studies on bovine thyroidal purine nucleoside phosphorylase

Abstract 1. 1. Bovine thyroid purine nucleoside phosphorylase shows negative cooperativity in the presence of phosphate with a Hill coefficient of 0.56 which is influenced by pH. 2. 2. Initial velocity and product inhibition studies show the enzyme reaction to be a Bi-Bi ordered mechanism. 3. 3. Of the substrates, phosphate binds first followed by the nucleoside and the release of nucleobase precedes the release of ribose-1-phosphate for the products.