[The correcting role of autonomous 3'-->5' exonucleases in mammalian multienzyme DNA polymerase complexes].
A study was made of the correcting role of autonomous 3'-->5' exonucleases (AE) contained in multienzyme DNA polymerase complexes of rat hepatocytes or calf thymocytes. DNA was synthesized on phage psi X174 amber3 or M13mp2 primer-templates, and used to transfect Escherichia coli spheroplasts. Frequencies were estimated for direct and reverse mutations resulting from mistakes made in the course of in vitro DNA synthesis. The mistake rate of the hepatocytic complex was estimated at 3 x 10(-6) with equimolar dNTP, and increased tenfold when proteins accounting for 70% of the total 3'-->5' exonuclease activity of the complex were removed. The fidelity of DNA synthesis was completely restored in the presence of exogenous AE (epsilon subunit of E. coli DNA polymerase III). Nuclear (Pol delta n) and cytosolic (Pol delta c) forms of DNA polymerase delta were isolated from calf thymocytes. The former was shown to contain an AE (TREX2) absent from the latter. As compared with Pol delta c, Pol delta n had a 20-fold higher exo/pol ratio and allowed 4-5 times higher fidelity of DNA synthesis. The mistake rate of DNA polymerase complexes changed when dNTP were used in nonequimolar amounts.
Shevelev
I V
IV
Institute of Nuclear Physics, Russian Academy of Sciences, Gatchina, Leningrad Oblast, 188300 Russia.
Beliakova
N V
NV
Kravetskaia
T P
TP
Krutiakov
V M
VM
D000818:Animals; D001435:Bacteriophages / Q000235:genetics; D001671:Biochemistry / Q000379:methods; D002417:Cattle; D002467:Cell Nucleus / Q000201:enzymology / Q000235:genetics; D003593:Cytoplasm / Q000201:enzymology / Q000235:genetics; D004247:DNA / Q000096:biosynthesis; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D043211:Exodeoxyribonuclease V; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D022781:Hepatocytes / Q000201:enzymology; D009097:Multienzyme Complexes / Q000378:metabolism*; D051381:Rats; D013950:Thymus Gland / Q000201:enzymology
D009097:Multienzyme Complexes; D004247:DNA; D004259:DNA-Directed DNA Polymerase; D005090:Exodeoxyribonucleases; D043211:Exodeoxyribonuclease V