Acta biochimica et biophysica; Academiae Scientiarum Hungaricae最新文献

Binding of beta-adrenergic receptor by labelled dihydroalprenolol was inhibited in the presence of GTP or GppNHp in rat brain particulate fractions. The inhibition was suspended when the particulate fraction was pretreated at 30 degrees C for 20 minutes and washed 3 times. It is concluded that the inhibition was due to endogenous bound catecholamines.

Two forms of deoxythymidine kinase were separated from human tonsillar lymphocytes on DEAE-Sephadex column (peak 1 and peak 2 isoenzymes) and identified by gel electrophoresis. Both isoenzymes were found in freshly prepared lymphocytes as well as in lymphocytes cultured in the presence or absence of phytohaemagglutinin, but freshly prepared and phytohaemagglutinin-stimulated cells contained higher activity of peak 1 isoenzyme than the cultured, non-stimulated ones. Kinetic properties and feed-back inhibition of the two separated isoenzymes were compared. The apparent Km values of isoenzyme 1 and 2 were 4 microM and 5 microM for thymidine and were 0.15 mM and 0.12 mM for ATP, respectively. Isoenzyme 1 was found to be more sensitive to heat denaturation at 55 degrees C, and to feedback inhibition by dTTP than isoenzyme 2. On the contrary, isoenzyme 1 was more resistant to dCTP inhibition than the other one. It is concluded that the isoenzyme 1 activity correlates with the synthesis of DNA, while the activity of isoenzyme 2 seems to be constant. The high activity of isoenzyme 1 in freshly isolated tonsil lymphocytes supports their active proliferation in the organ.

During isolation pancreatic elastase is accompanied by a minor component, elastomucoproteinase (EMPase), which can be separated in pure state by anion-exchange chromatography. EMPase exhibits both proteolytic and mucolytic activities. Applying powdered aortic preparations as a substrate, the enzyme can split off carbohydrate moieties, but it can also cleave peptide bonds as a proteinase. Assaying with tripeptide-p-nitroanilide substrates, the enzyme can decompose only those substrates which contain Phe or Tyr at the C-terminal. It appeared that the binding of Bz-Ala-Gly-Phe-pNA was the best, while the hydrolysis of Bz-Ala-Pro-Tyr-pNA was the fastest. Proteolytic activity of EMPase can be completely inhibited with phenylmethanesulphonyl fluoride, whereas its mucolytic activity only by 20%. The two activities may be located at two separate sites. The enzyme seemed to be composed of a single polypeptide chain by SDS-gel electrophoresis in the presence of urea and beta-mercapto-ethanol.

Endothelial cells, under normal conditions, possess antithrombotic nature, whereas during damage, various components of the cell may initiate blood coagulation. These functions are greatly influenced and controlled by thrombin and plasmin through their direct actions or their formation in the blood coagulation-fibrinolytic system.

By using recombinant DNA techniques an artificial operon was constructed that codes for two fusion proteins under the control of the beta-lactamase promoter of plasmid pBR322. The two proteins are: 1. beta-lactamase-trp B (43 kd) 2. trpA-beta-galactosidase (120 kd). Frameshift mutations in the N-terminal region of the first gene resulted in a dramatic reduction in the synthesis of the protein coded by the second gene. This strong polar effect could not be accounted for by correspondingly lower level of the distal region of the messenger RNA, only by "translational coupling" due to the overlap of the termination codon of the first gene with the initiation codon of the second gene. It was concluded that the strong "translational coupling" observed in this artificial operon can be generally used to ensure coordinated high-level synthesis of proteins in operons constructed by recombinant DNA techniques.

The heat sensitivity and stability of a protein should be characterized by the Arrhenius parameters of the irreversible denaturation. The terms "thermostability" or "activation enthalpy" and "activation entropy" of denaturation as "thermodynamic parameters of protein stability" are misleading. The terms "measure of heat sensitivity" and "measure of stability" should be used as comparative data for protein structure.

Thrombin and plasmin are inactivated by antithrombin III at different rates. Heparin, at a catalytic amount, increases primarily the inactivation rate of thrombin. The two proteinases compete for heparin, i.e. heparin is preferably attached by thrombin. Furthermore, fibrinogen as well as fibrin protect plasmin against inactivation by antithrombin III. However, at high concentration of heparin, plasmin inactivation by antithrombin III is accelerated even in the presence of fibrinogen or fibrin.