scholarly journals Inhibition of murine leukaemia virus reverse transcriptase by 2-halogenated polyadenylic acids

1982 ◽  
Vol 203 (3) ◽  
pp. 755-760 ◽  
Author(s):  
T Fukui ◽  
E De Clercq
Keyword(s):  
Reverse Transcriptase ◽  
Dna Polymerase ◽  
Reverse Transcriptase Activity ◽  
Leukaemia Virus ◽  
Transcriptase Activity ◽  
Murine Leukaemia Virus ◽  
Polyadenylic Acid ◽  
Moloney Murine Leukaemia Virus

Several new analogues of polyadenylic acid [(A)n], i.e. poly(2-fluoroadenylic acid) [(fl2A)n], poly(2-chloroadenylic acid [(cl2A)n], poly(2-bromoadenylic acid) [(br2A)n] and poly(2-iodoadenylic acid) [(io2A)n] have been synthesized and evaluated for their effects on the RNA-directed DNA polymerase (reverse transcriptase) activity of Moloney murine leukaemia virus. All (A)n analogues were found to be potent inhibitors of reverse transcriptase, the order of (decreasing) potency being (fl2A)n greater than (io2A)n greater than (br2A)n greater than (cl2A)n. For all four (A)n analogues the inhibition of reverse transcriptase was competitive with respect to the template-primer. (A)n . oligo(dT). The K1 values were 0.02 microgram/ml for (fl2A)n, 0.1 microgram/ml for (io2A)n, 0.5 microgram/ml for (br2A)n and 8 microgram/ml for (cl2A)n. With a Ki of 0.02 microgram/ml (approx. 0.04 microM), (fl2A)n can be considered as one of the most, if not the most, potent polynucleotide inhibitor of reverse transcriptase that has been described so far.

scholarly journals Sulphydryl groups in the template-primer-binding domain of murine leukaemia virus reverse transcriptase. Identification and functional analysis of cysteine-90

1993 ◽  
Vol 296 (3) ◽  
pp. 577-583 ◽  
Author(s):  
S Basu ◽  
A Basu ◽  
M J Modak
Keyword(s):  
Reverse Transcriptase ◽  
Dna Polymerase ◽  
Structural Integrity ◽  
Peptide Mapping ◽  
Site Directed Mutagenesis ◽  
Heat Inactivation ◽  
Composition Analysis ◽  
Kinetic Measurements ◽  
Leukaemia Virus ◽  
Murine Leukaemia Virus

Treatment of murine leukaemia virus reverse transcriptase with benzophenone 4-maleimide inactivates DNA polymerase activity, but has no effect on the RNAase H function. Kinetic measurements indicated that benzophenone 4-maleimide is a competitive inhibitor with respect to template-primer binding, but is non-competitive with respect to dNTP binding. Enzyme modified with benzophenone 4-maleimide cannot bind template-primer or primer alone, as judged by u.v.-mediated cross-linking of radiolabelled substrates. Of the eight cysteine residues in murine leukaemia virus reverse transcriptase, only two were modified by benzophenone 4-maleimide, which were identified as Cys-90 and Cys-310 by comparative tryptic-peptide mapping and amino acid composition analysis. Inclusion of template-primer or primer alone in the modification mixture protected only Cys-90 from modification by benzophenone 4-maleimide. To investigate the role of Cys-90 in detail, we converted it to alanine by site-directed mutagenesis. The mutant enzyme, however, exhibited no loss either of DNA polymerase or of RNAase H activity. These results indicate that Cys-90 is located in a domain of murine leukaemia virus reverse transcriptase that binds template-primer, but may not have a direct role in the enzymic function of the enzyme. Ala-90 mutant murine leukaemia virus reverse transcriptase is at least 10-fold more susceptible to heat inactivation than is the wild-type enzyme, which suggests that Cys-90 in murine leukaemia virus reverse transcriptase may play a role in maintaining structural integrity.

scholarly journals Complexing reverse transcriptase with polyspermine-ribonuclease

1980 ◽  
Vol 189 (1) ◽  
pp. 89-93
Author(s):  
A K Bandyopadhyay ◽  
D Wang ◽  
C C Levy
Keyword(s):  
Complex Formation ◽  
Reverse Transcriptase ◽  
Reverse Transcriptase Activity ◽  
Favourable Condition ◽  
Molar Ratio ◽  
Leukaemia Virus ◽  
Transcriptase Activity

Polyspermine-ribonuclease (Mr approximately 17 000) and the enzyme transcriptase from Rauscher-leukaemia virus (Mr approximately 70 000) form a complex Mr approx. 160 000) such that the molar ratio of polyspermine-ribonuclease to reverse transcriptase is 5:1. The most favourable condition for complex-formation is in a solution consisting of 0.01 M-Tris/HCl buffer, pH 7.5, 0.25 M-KCl and 1 mM-Mn2+ at 37 degrees C. The association of the two enzymes retains full RNAase activity, but reverse-transcriptase activity is completely inhibited when ribonuclease-sensitive polymers such as (dG)12 x (rC)n or viral 70S RNA are used as primer templates.

scholarly journals Limited reverse transcriptase activity of phi29 DNA polymerase

2018 ◽  
Vol 46 (7) ◽  
pp. 3625-3632 ◽  
Author(s):  
Tomasz Krzywkowski ◽  
Malte Kühnemund ◽  
Di Wu ◽  
Mats Nilsson
Keyword(s):  
Reverse Transcriptase ◽  
Dna Polymerase ◽  
Reverse Transcriptase Activity ◽  
Transcriptase Activity ◽  
Phi29 Dna Polymerase
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