Study of 2'-phosphodiesterase activity in cultured NIH 3T3 cells during activation of cAMP-dependent phosphorylation

A rapid and transient decrease in 2'-phosphodiesterase activity in NIH 3T3 mouse cells was observed after adrenaline addition. The decrease of activity was accompanied by an elevation of intracellular cAMP level. The 2'-phosphodiesterase activity changed similarly when cells sink deeper into the resting state. In the latter case, the fall of the enzyme activity was correlated with elevation of the activity of cAMP-dependent proteinkinase and, moreover, a considerable increase of the intracellular level of 2',5'-oligoadenylate was observed. Phosphorylation of proteins by cAMP-dependent proteinkinase in the cell lysate also produced a pronounced drop of 2'-phosphodiesterase activity. Exogenous 2',5'-oligo (A) treatment of the cells resulted in the rise of 2'-phosphodiesterase activity; actinomycin D prevented this effect. The data presented suggest the involvement of two different mechanisms in regulation of 2'-phosphodiesterase activity: cAMP-dependent phosphorylation and induction of 2'-phosphodiesterase by 2',5'-oligoadenylate.     

Mechanism of action of cAMP-dependent protein kinase. V. Free energy spectra

The catalytic subunit of cAMP-dependent protein kinase from pig brain was shown to catalyse an isotope exchange reaction ATP in equilibrium with ADP. The kinetic parameters of the exchange were determined. The enzyme can also use GTP as the donor substrate; phosphotransferase and "GTPase" reactions were investigated. Based on the kinetic data obtained in this and in the previous paper the free energy profiles of protein kinase catalysed reactions are discussed.     

Dipyridamole-induced interferon production in mouse peritoneal leukocytes

The in vitro explanted mouse peritoneal leukocytes were used for the optimization of the dipyridamole-induced interferon production. After 90-120 min incubation of cells with 30-100 microM dipyridamole, the production of interferon reached 6.4 X 10(4) IU/ml. It was demonstrated that the interferon production phase is preceded by the dipyridamole-dependent increase in cAMP concentration. The possibility of cAMP involvement in the mechanism of interferon production is being discussed.     

Localization of histone H1 in chromatin. Cross-linking of the N- and C-terminal halves of the molecule with bifunctional reagents

Mutual arrangement of histone H1 molecules was studied in calf thymus nuclei, extended chromatin and chromatin, isolated and kept in 8 M urea. Histone H1 dimers crosslinked with methyl 4-mercaptobutyrimidate were digested with chymotrypsin and crosslinked fragments obtained were analysed by diagonal gel electrophoresis. In all chromatins tested the N- and C-terminal parts of the H1 molecules were crosslinked in all possible combinations, i.e. C-C, C-N and N-N. These and related data obtained earlier indicate, that the proximity of histone H1 molecules in chromatin is determined by the structure of nucleosomal chain itself and not by chromatin superstructure. The results also suggest that the H1A and H1B subfractions of histone H1 are interspersed in extended nucleosomal chains.     

Study on the role of SH-groups in the activity of muscle pyruvate dehydrogenase

The kinetics of inactivation of the pyruvate dehydrogenase component of the pigeon breast muscle pyruvate dehydrogenase complex in the presence of 5,5'-dithiobis (2-nitrobenzoate) is biphasic. The rate constants for the fast and slow phases of the inactivation reaction are close to those for modification of two classes of SH-groups differing in their reactivities towards the inhibitor. The reaction order with respect to the inhibitor concentration suggests that the two distinct SH-groups are essential for the enzyme activity. Modification of these SH-groups results in inhibition of the overall activity of the pyruvate dehydrogenase complex and of the 2-hydroxyethyl thiamine pyrophosphate - acceptor oxidoreductase activity of its decarboxylating component. Thiamine pyrophosphate exerts a protective effect on the enzyme only at the slow phase of the enzyme inactivation and SH-modification. As a result of interaction between the holoenzyme and pyruvate (or apoenzyme and 2-hydroxyethyl thiamine pyrophosphate) the rate of the enzyme inactivation is increased. This is associated with masking of non-essential SH-groups and with an increase of the accessibility of two essential SH-groups to the inhibitor. The data obtained suggest the interrelationship between the essential SH-groups and the 2-hydroxyethyl thiamine pyrophosphate-acceptor oxidoreductase activity of pyruvate dehydrogenase.     

Peristaltic flow in tubes

The study is concerned with the analysis of two flow domains of peristaltic motion in tubes. In the first analysis the wall disturbance wavelength is much larger than the average tube radius. There is a simple algebraic relation between the average flow rate and pressure differential across a wavelength. In the second analysis the disturbance wavelength may be as small as the average radius. A numerical technique may be used to determine the relation between average flow rate and pressure differential across a wavelength.