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M J Modak

Publications and source records attributed to M J Modak.

At least 109 records · Page 6Linked to original sources
PubMed1976

Observations on the pyridoxal 5'-phosphate inhibition of DNA polymerases.

Pyridoxal 5'-phosphate at concentrations greater than 0.5 mM inhibits polymerization of deoxynucleoside triphosphate catalyzed by a variety of DNA polymerases. The requirement for a phosphate as well as aldehyde moiety of pyridoxal phosphate for inhibition to occur is clearly shown by the fact that neither pyridoxal nor pyridoxamine phosphate are effective inhibitors. Since the addition of nonenzyme protein or increasing the amount of template primer exerted no protective effect, there appears to be specific affinity between pyridoxal phosphate and polymerase protein. The deoxynucleoside triphosphates, however, could reverse the inhibition. The binding of pyridoxal 5'-phosphate to enzyme appears to be mediated through classical Schiff base formation between the pyridoxal phosphate and the free amino group(s) present at the active site of the polymerase protein. Kinetic studies indicate that inhibition by pyridoxal phosphate is competitive with respect to substrate deoxynucleoside triphosphate(s).

Animals↗
PubMed1976

Observations on template-specific conditions for DNA synthesis by avian myeloblastosis virus DNA polymerase.

The effects of Mg++, Mn++, and KCl addition, individually and in combination, on the rate of DNA- and RNA-primed DNA synthesis by avian myeloblastosis virus DNA polymerase (reverse transcriptase) using a variety of natural and synthetic template-primer combinations were examined. Optimal divalent cation concentrations were found to vary by as much as 10-fold depending upon the template-primer used to direct synthesis. Addition of KCl to reaction mixtures containing optimal divalent cation concentrations produced stimulation or inhibition of DNA synthesis which was also template-specific. DNA synthesis on the modified template poly (2'-0-methylcytidylate) was uniquely stimulated by combinations of divalent cations. With Mg++ as divalent cation, deviations from classical Michaelis-Menten kinetics of substrate saturation were observed with all template-primers tested.

Avian Leukosis Virus↗
PubMed1974

Evidence for allosterism in in vitro DNA synthesis on RNA templates.

Hemoglobin mRNA and (rA)(n).(dT)(10) have been used as primer-templates in a kinetic study of DNA synthesis with Escherichia coli DNA polymerase I (DNA nucleotidyl transferase, EC 2.7.7.7) and Mason-Pfizer monkey virus reverse transcriptase (RNA-directed DNA polymerase). The rate versus enzyme concentration curve is sigmoidal and is consistent with a cooperative phenomenon. The results could be interpreted in terms of the formation of an active complex containing enzyme dimers (or oligomers) on the primer-template. We have also observed sigmoidal kinetics in rate versus deoxynucleotide triphosphate concentration. These results are consistent with an allosteric mechanism in which the triphosphates act as both modifiers and DNA precursors. In the critical range, a 6- to 8-fold increase in both enzyme and triphosphate concentrations can lead to a 1500-fold increase in the rate of synthesis on an RNA template. Thus, small changes in enzyme and precursor concentrations could play a regulatory role in vivo.

Adenosine Triphosphate↗
PubMed1974

Purification of avian myeloblastosis virus DNA polymerase by affinity chromatography on polycytidylate-agarose.

Polycytidylic acid [poly(rC)] covalently linked to cyanogen bromide-activated agarose is an effective affinity matrix for the RNA-dependent DNA polymerase from avian myeloblastosis virus. Poly(rC)-agarose is capable of binding large quantities of avian myeloblastosis DNA polymerase, which is then eluted by using a linear KCl gradient of increasing concentration. The DNA polymerase isolated from crude, detergent-disrupted virions by a single pass through columns of poly(rC)-agarose appears nearly homogeneous (approximately 90% pure) as determined by sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis. Complete recovery of input enzymatic activity was obtained. Results suggest that polyribonucleotide columns may provide a high-yield, rapid method for the purification of oncornaviral DNA polymerase.

Avian Leukosis Virus↗