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DNA synthesis primed by mononucleotides (de novo synthesis) catalyzed by HIV‐1 reverse transcriptase: tRNA Lys,3 activation
Author(s) -
Zakharova Ol'ga D.,
Tarrago-Litvak Laura,
Fournier Michel,
Litvak Simon,
Nevinsky Georgyi A.
Publication year - 1995
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(95)01056-k
Subject(s) - primer (cosmetics) , dna synthesis , oligonucleotide , reverse transcriptase , transfer rna , nucleotide , de novo synthesis , chemistry , dna , polymerase , enzyme , stereochemistry , dna polymerase , biochemistry , protein biosynthesis , microbiology and biotechnology , chain termination , biology , polymerization , rna , gene , organic chemistry , radical polymerization , polymer
HIV‐1 RT is able to catalyze DNA synthesis starting from mononucleotides used both as minimal primers and as nucleotide substrates (de novo synthesis) in the presence of a complementary template. The rate of this process is rather slow when compared to the polymerization primed by an oligonucleotide. The addition of tRNA Lys,3 to this system increased the de novo synthesis rate by 2‐fold. Addition of low concentrations of agents able to modify protein conformation, such as urea, dimethylsulfoxide and Triton X‐100, can activate the de novo synthesis by a factor 2 to 5. A dramatic synergy is observed in the presence of the three compounds since the stimulating effect of tRNA increases 10–15 times. These results suggest that compounds activating RT are able to induce a conformational change of the enzyme which results in a higher specific activity. Primer tRNA seems to play an important role in HIV‐1 RT modification(s) leading to a polymerase having a higher affinity for the primer or the dTTP, but not for the template. The specificity of RT for the template is not influenced by changes in the kinetics or in the thermodynamic parameters of the polymerization reaction.