Changes In Relative Activity Research Articles

The effects of thyroid hormones on monoamine oxidase in the rat heart

Abstract The administration of thyroxine to young male rats produced an increase in the specific activity of their cardiac monoamine oxidase (MAO). A reduction in the circulating concentrations of thyroid hormones, brought about by 2-thiouracil, led to a decrease. The relative change in activity produced was greater with tyramine than with benzylamine as substrate. By following the time-course of the return of enzyme activity, with tyramine as substrate, after a single injection of pargyline in vivo, it was concluded that both excess and lack of thyroid hormones cause their effects on MAO activity by changing the rate of synthesis of the enzyme and not its degradation rate constant. The degradation rate constant did change with the age of the animal. The MAO activity, which increased towards tyramine as substrate in hyperthyroid rat hearts, behaved in the same way as that of controls to heat treatment, irreversible inhibition by pargyline or by clorgyline and also in Km determinations. The pattern for benzylamine oxidation was similar, except for the effect of the inhibitor clorgyline which shifted the plateau region of the double sigmoid inhibition curve significantly using enzyme from hyperthyroid rat hearts. The plateau region was also shown to be affected by the age of the animal. The possibility is discussed that the increased cardiac MAO activity produced by thyroid hormones and by the growth of the animal is mediated by that form of the enzyme primarily responsible for the oxidation of tyramine. Mixed substrate experiments suggested that tyramine oxidation could be inhibited competitively by benzylamine.

Separation and Partial Characterization of Two Ribonucleic Acid Polymerases from Pea Seedlings

Two DNA-dependent RNA polymerases (ribonucleoside triphosphate:RNA nucleotidyl transferase, EC 2.7.7.6) have been isolated from pea (Pisum sativum) seedlings. The enzymes were solubilized by sonication in high salt buffer and were separated by chromatography on diethylaminoethyl cellulose using a linear salt gradient. Polymerase I eluted at 0.10 m (NH(4))(2)SO(4), accounted for about 10% of the recovered activity and was completely insensitive to alpha-amanitin. Polymerase II eluted at 0.14 m (NH(4))(2)SO(4), accounted for the remaining 90% of recovered activity and was strongly inhibited by alpha-amanitin. Both enzymes preferred denatured to native DNA as template, both showed an absolute requirement of divalent cation, and both were sensitive to the ionic strength of the assay medium. The developing pea seedling seems a promising system for studies of possible changes in relative activities and roles of multiple RNA polymerases during eukaryotic development.

Changes in enzymic activity and distribution of acid-soluble, amino acid-nitrogen in soil during nitrogen immobilization and mineralization

During a period of immobilization of nitrate- 15N and mineralization of organic N in a sandy-loam, changes were recorded in: (a) the concentration of an added carbon source, glucose- 14C: (b) evolution of 14CO 2: (c) bacterial populations; (d) distribution and concentration of newly-synthesized, acid-soluble, amino acid- 15N; and (e) distribution and activities of several oxidative and hydrolytic enzyme systems. Added glucose- 14C was rapidly metabolized by the soil microflora. After 1.5 day's incubation, when only 3.6 per cent of the added glucose was present, 68 per cent of the 14C remained in the soil-microbial system. During this period there was a marked increase in viable bacterial numbers and an almost complete immobilization of nitrate- 15N. On continued incubation, microbial metabolites were oxidized at decreasing rates, the more rapid phase corresponding to a period of net decline in the viable bacterial population. Soil was fractionated by a relatively mild procedure into components containing: (a) extractable proteins; (b) extractable amino acids and peptides; (c) particulate material containing microbial cells, cell debris and material bound to larger soil particles; and (d) microbial metabolites mainly bound to soil colloids. Although the total, acid-soluble, amino acid- 15N remained relatively constant for about 50 days, there were marked changes in their concentration in different fractions, especially in the extracts and in the fraction containing fine colloidal material. However, the relatively large decline in labelled, acid-soluble, amino acid- 15N occurred during the active phase of oxidation of microbial metabolites when little net mineralization of labelled compounds occurred. Increases in enzymic activities generally coincided with increased viable bacterial populations although there were some exceptions, notably casein and benzoyl arginine amide-hydrolysing enzymes. The stabilities of the newly-formed enzymes varied markedly. The greatest relative changes in activity occurred with the casein-hydrolysing enzymes. Their activity reached a maximal value after the main flush of bacterial growth, was short-lived and was to a large extent extractable. The formation and disappearance of this extracellular proteolytic activity coincided approximately with that of a secondary peak of extractable, newly-synthesized, protein- 15N. In general however, changes in enzymic activity could not be identified with changes of protein- 15N concentrations of the different fractions.

Кoличecтвeннoe oпpeдeлeниe cпeцифичecкич ингибитopoв пoлимepизaции фибpинa. Eдиницы тopмoзящeй aктивнocти

Fibrinogen and its degradation products are to be regarded as specific inhibitors of fibrin polymerization because of the protein-protein recognizing manner of their action. Some effects of specific inhibitors, e.g. prolongation of clotting time are easily measurable, yet, a proper quantitative presentation of the inhibitory activity is a separate problem requiring consideration. No series of direct measurements of inhibitory effects can adequately reflect the relative activities of inhibitors in question as the inhibitor concentration - effect dependence is non-linear. In Figure 1 curves for fibrinogen and fragment D are shown. Ordinate - relative prolongation of pure fibrin monomer clotting, abscissa - inhibitor concentration. The curves suggest some cooperativity in fibrin-inhibitor interactions. Dotted lines indicate a striking change in relative activity of the two inhibitors due to altering the concentration chosen for the activity-comparison test. Thus, the mere values of the inhibitory effects can give hardly more than semi-quantitative hints on the intrinsic inhibitory activities. To gain true values we made resort to the estimated weight quantities of inhibitors producing the same effect. As these quantities are obviously inversely proportional to the specific activity values of the respective inhibitors they immediately provide the information wanted. An arbitrary standard was chosen: 10-fold retardation of fibrin monomer clotting at definite medium conditions. So it became possible to express activities of specific inhibitors in terms of special units. In mixtures of inhibitors the overall activity (the total amount of activity units) was found equal or near to the sum of unit contents of all the components present. Consequently, the behaviour of components is virtually additive (Tables 2 and 3). This additivity means that the effects of individual components enhance each other in the same co-operative manner as the effectiveness of a single inhibitor is raised by its concentration increase (Figure 1). The arithmetical sum of separately exerted effects of components generally proved far below the actual effect of the corresponding mixture. This difference, as expected theoretically, was maximal if the components were represented in equivalent amounts, and it was less for a two-component system than for a three-component one. These results warrant the adequacy of the new method. Following determinations were carried out with this method. In fibrinogen solutions subjected to tryptic hydrolysis the activity unit content increased by about 300 per cent under optimal conditions, i.e. in the presence of Ca 2+ and at 1:2250 trypsin-fibrinogen weight ratio. Specific activities of fibrinogen and purified fragment D were found to amount to 1.4 and 10.9 units per mg respectively. Throughout our work on fragment D isolation from tryptic fibrinogen digests activity yields were determined to check the procedure under trial.

Enzyme Activities in Polarized Cell Membranes

The theoretical pH dependence of enzyme activities in membranes of low dielectric constant is estimated. It is shown that in biological membranes some types of enzymes may attain a limiting pH sensitivity such that an increment of only 0.2 pH unit (sufficient to induce action potentials in squid axons) causes a relative activity change of over 25%. The transients of enzyme activity generated by membrane depolarization and by pH increments in the bathing solution are discussed in relation to the transients of nervous excitation.