Differential inhibitors of DNA polymerases alpha and delta

DNA polymerases alpha and delta from bone marrow are similar in many respects, the major known difference being the exonuclease activity of delta. Differential inhibitors of alpha and delta have been sought to assist in their functional and physical separation. Butylphenyl deoxyguanosine triphosphate is one. It effectively inhibits alpha at less than 1 microM concentration, whereas more than 100 microM is required to similarly inhibit delta. Another is the monoclonal antibody, SJK 132-20, which neutralizes the polymerase activity of alpha but not delta. These differential inhibitors further define alpha and delta as separate categories of eukaryotic DNA polymerase and promise to facilitate the study of both.     

Photoconversion of 7-dehydrocholesterol to vitamin D3 in synthetic phospholipid bilayers

The incorporation of 7-dehydrocholesterol into synthetic phospholipid bilayers altered the distribution of products after photolysis. In liposomes, the relative amounts of 7-dehydrocholesterol and lumisterol were elevated, and tachysterol was reduced from the levels observed in hexane solution. Z to E isomerization of the previtamin to tachysterol is favored in organic solvents. The inhibition of this process is evidence that an ordered lipid matrix places a new constraint on the conformation of the ring B fission product--one in which the configuration is favorable for a return to a cyclized diene. Further, rate enhancements of up to 15-fold were observed for the thermal isomerization of the previtamin to vitamin D3 in liposomes. The free energies of activation for the reaction at 25 degrees C were reduced by 1.3-1.5 kcal/mol in the bilayer environment compared to that of hexane. As this reaction involves the concerted transfer of a hydrogen via a cyclic intermediate, it provides additional evidence for membrane stabilization of an all-cis conformation of the previtamin. Photoproduct ratios were also studied for 7-dehydrocholesterol adsorbed to a variety of solid supports. That nonspecific interactions of 7-dehydrocholesterol with lipid can influence product formation may have important implications with respect to the mechanism of vitamin D3 biosynthesis.     

The reduction of alpha-ketocarboxylic acids and alpha-dicarbonyl compounds with titanium(III) chloride

Thirteen model alpha-ketocarboxylic acids and alpha-dicarbonyl compounds have been observed to be reduced by titanium(III) chloride. The products of these reactions were shown by melting (boiling) points, mixed melting points, derivatives, refractive indices, infrared, and NMR comparisons with authentic compounds or literature information to be the corresponding alpha-hydroxycarboxylic acids or alpha-hydroxyketones.     

Demonstration of the presence of G-proteins in hepatic microsomal fraction

The presence of G-proteins in isolated hepatic microsomal vesicles is demonstrated. The G-proteins were identified by their capacity to be ADP-ribosylated by cholera and pertussis toxins. Cholera toxin identified 42 and 45 kDa proteins, corresponding to alpha s-1 and alpha s-2, respectively. Pertussis toxin identified a 40 kDa protein corresponding to alpha i. The microsomal G-proteins are identical to the corresponding G proteins of the plasma membrane, but are present in different proportions; the microsomes have considerably less alpha s proteins than the plasma membrane.     

Cyclic nucleotide alterations in mixed leukocyte reaction: a preliminary report

Endogenous cyclic adenosine and guanosine monophosphate (cAMP, cGMP) levels were studied in human peripheral blood lymphocytes during mixed leukocyte reactions (MLR). cAMP level was consistently elevated in one-way MLR, with good correlation to 3H-thymidine uptake in these reactions. In contrast, cGMP level was practically unchanged. Irradiation of reacting cell populations resulted in inhibition of cyclic nucleotide phosphodiesterase (PDE) activity. These results suggest that metabolic alterations in cAMP may be associated with immune reactions of cellular recognition.     

Regulation of lung endothelin content by the glucocorticosteroid budesonide.

Intratracheal instillation of Sephadex beads induced a long-lasting inflammation in the rat lung as seen by an increase in lung weights. Repeated instillation enhanced this reaction and increased lung endothelin-1 content 3.5 times. Budesonide given s.c. abolished these effects and even reduced basal endothelin-1 content by 72%. The tissue content of the sensory neuropeptide neurokinin A were unaffected by both treatments. Endothelin has been proposed to play a part in the pathogenesis of bronchial asthma. If it is so, the ability of budesonide to reduce endothelin-1 content could thus be added to the list of beneficial effects of glucocorticosteroids in these conditions.     

Effects of troponin I phosphorylation on conformational exchange in the regulatory domain of cardiac troponin C.

Conformational exchange has been demonstrated within the regulatory domain of calcium-saturated cardiac troponin C when bound to the NH2-terminal domain of cardiac troponin I-(1-80), and cardiac troponin I-(1-80)DD, having serine residues 23 and 24 mutated to aspartate to mimic the phosphorylated form of the protein. Binding of cardiac troponin I-(1-80) decreases conformational exchange for residues 29, 32, and 34. Comparison of average transverse cross correlation rates show that both the NH2- and COOH-terminal domains of cardiac troponin C tumble with similar correlation times when bound to cardiac troponin I-(1-80). In contrast, the NH2- and COOH-terminal domains in free cardiac troponin C and cardiac troponin C bound cardiac troponin I-(1-80)DD tumble independently. These results suggest that the nonphosphorylated cardiac specific NH2 terminus of cardiac troponin I interacts with the NH2-terminal domain of cardiac troponin C.