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Monofunctional DNA-platinum(II) adducts block frequently DNA polymerases
1Institut für Biophysik und physikalische Biochemie, Universität Regensburg, Germany.
Abstract:
The question of whether monofunctional DNA platinum(II) adducts block synthesis of DNA by purified DNA polymerases of different types and origin has been investigated by comparing the time dependence of synthesis arrest and of DNA adduct formation. Activated salmon testis DNA is used as a suitable substrate for DNA synthesis allowing to probe inhibition by platinum(II) monoadducts for the variety of inherent template-primers. Reaction amplitudes are related to defined mixtures of dichloro and chloroaqua platinum(II) complexes. It is found that (i) all investigated DNA polymerases seem arrested (100% efficiency) at bifunctional DNA adducts. (ii) human DNA polymerase beta bypasses most of the monofunctional lesions of the three platinum(II) complexes investigated. (iii) Klenow fragment is blocked by monoadducts with increasing efficiency in the order cis-diamminechloroaquaplatinum(II) (0%) less than meso-[1,2-bis(2,6- dichloro-4-hydroxyphenyl)ethylenediamine] chloroaquaplatinum(II) (50%) less than trans-diamminechloro-aquaplatinum(II) (75%). (iv) Escherichia coli DNA polymerase I, Thermus aquaticus DNA polymerase, Physarum polycephalum DNA polymerase alpha, and calf thymus DNA polymerase alpha appear to be arrested by monoadducts. According to these examples, blocking efficiencies depend on the cis/trans-stereogeometry of fixation of the carrier ligands at platinum(II) residues, on the size/chemical nature of the platin(II) carrier ligand and on the type/origin of DNA polymerase.
Insights
Monofunctional platinum(II) DNA adducts block DNA synthesis by most polymerases, but human DNA polymerase beta bypasses these lesions. Blocking efficiency depends on platinum complex structure and DNA polymerase type.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Platinum(II) complexes are used in cancer chemotherapy.
- DNA adducts can interfere with DNA replication and transcription.
- Understanding polymerase bypass of DNA adducts is crucial for drug development.
Purpose of the Study:
- To investigate if monofunctional DNA platinum(II) adducts block DNA synthesis by various DNA polymerases.
- To compare the inhibition efficiency of different platinum(II) complexes on DNA synthesis.
- To determine factors influencing DNA polymerase arrest at platinum(II) adducts.
Main Methods:
- Utilized activated salmon testis DNA as a substrate for DNA synthesis.
- Compared time-dependent DNA synthesis arrest with DNA adduct formation.
- Investigated reactions involving dichloro and chloroaqua platinum(II) complexes.
- Tested purified DNA polymerases from different species and origins.
Main Results:
- All tested DNA polymerases were 100% arrested by bifunctional DNA adducts.
- Human DNA polymerase beta efficiently bypassed most monofunctional platinum(II) lesions.
- Klenow fragment showed increasing blockage by monoadducts: cis-diamminechloroaquaplatinum(II) (0%) < meso-[1,2-bis(2,6-dichloro-4-hydroxyphenyl)ethylenediamine] chloroaquaplatinum(II) (50%) < trans-diamminechloro-aquaplatinum(II) (75%).
- Escherichia coli DNA polymerase I, Thermus aquaticus DNA polymerase, Physarum polycephalum DNA polymerase alpha, and calf thymus DNA polymerase alpha were arrested by monoadducts.
Conclusions:
- DNA polymerase arrest by monofunctional platinum(II) adducts varies significantly.
- Blocking efficiency is influenced by the stereochemistry (cis/trans) of platinum(II) complexes.
- The size and chemical nature of the platinum(II) carrier ligand impact polymerase inhibition.
- DNA polymerase type and origin are critical determinants of adduct bypass or arrest.