Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein
O6-Alkylguanine-DNA alkyltransferases (AGTs) are proteins responsible for the removal of mutagenic alkyl adducts at the O6-atom of guanine and O4-atom of thymine. In the current study we set out to understand the role of the Ser134 residue in the Escherichia coli AGT variant OGT on substrate discrim...
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MDPI AG
2017-11-01
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| Series: | Molecules |
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| Online Access: | https://www.mdpi.com/1420-3049/22/11/1948 |
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doaj-57129a90d2c542cfa3f6394bf5b13f072020-11-25T01:41:36ZengMDPI AGMolecules1420-30492017-11-012211194810.3390/molecules22111948molecules22111948Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT ProteinNadia M. Schoonhoven0Derek K. O’Flaherty1Francis P. McManus2Lauralicia Sacre3Anne M. Noronha4M. Judith Kornblatt5Christopher J. Wilds6Department of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaDepartment of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaDepartment of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaDepartment of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaDepartment of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaDepartment of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaDepartment of Chemistry and Biochemistry, Concordia University, Montréal, QC H4B 1R6, CanadaO6-Alkylguanine-DNA alkyltransferases (AGTs) are proteins responsible for the removal of mutagenic alkyl adducts at the O6-atom of guanine and O4-atom of thymine. In the current study we set out to understand the role of the Ser134 residue in the Escherichia coli AGT variant OGT on substrate discrimination. The S134P mutation in OGT increased the ability of the protein to repair both O6-adducts of guanine and O4-adducts of thymine. However, the S134P variant was unable, like wild-type OGT, to repair an interstrand cross-link (ICL) bridging two O6-atoms of guanine in a DNA duplex. When compared to the human AGT protein (hAGT), the S134P OGT variant displayed reduced activity towards O6-alkylation but a much broader substrate range for O4-alkylation damage reversal. The role of residue 134 in OGT is similar to its function in the human homolog, where Pro140 is crucial in conferring on hAGT the capability to repair large adducts at the O6-position of guanine. Finally, a method to generate a covalent conjugate between hAGT and a model nucleoside using a single-stranded oligonucleotide substrate is demonstrated.https://www.mdpi.com/1420-3049/22/11/1948Bioorganic moleculesmodified oligonucleotidesDNA repairsubstrate specificitymutagenesishomology modelingDNA interstrand cross-link |
| collection |
DOAJ |
| language |
English |
| format |
Article |
| sources |
DOAJ |
| author |
Nadia M. Schoonhoven Derek K. O’Flaherty Francis P. McManus Lauralicia Sacre Anne M. Noronha M. Judith Kornblatt Christopher J. Wilds |
| spellingShingle |
Nadia M. Schoonhoven Derek K. O’Flaherty Francis P. McManus Lauralicia Sacre Anne M. Noronha M. Judith Kornblatt Christopher J. Wilds Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein Molecules Bioorganic molecules modified oligonucleotides DNA repair substrate specificity mutagenesis homology modeling DNA interstrand cross-link |
| author_facet |
Nadia M. Schoonhoven Derek K. O’Flaherty Francis P. McManus Lauralicia Sacre Anne M. Noronha M. Judith Kornblatt Christopher J. Wilds |
| author_sort |
Nadia M. Schoonhoven |
| title |
Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein |
| title_short |
Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein |
| title_full |
Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein |
| title_fullStr |
Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein |
| title_full_unstemmed |
Altering Residue 134 Confers an Increased Substrate Range of Alkylated Nucleosides to the E. coli OGT Protein |
| title_sort |
altering residue 134 confers an increased substrate range of alkylated nucleosides to the e. coli ogt protein |
| publisher |
MDPI AG |
| series |
Molecules |
| issn |
1420-3049 |
| publishDate |
2017-11-01 |
| description |
O6-Alkylguanine-DNA alkyltransferases (AGTs) are proteins responsible for the removal of mutagenic alkyl adducts at the O6-atom of guanine and O4-atom of thymine. In the current study we set out to understand the role of the Ser134 residue in the Escherichia coli AGT variant OGT on substrate discrimination. The S134P mutation in OGT increased the ability of the protein to repair both O6-adducts of guanine and O4-adducts of thymine. However, the S134P variant was unable, like wild-type OGT, to repair an interstrand cross-link (ICL) bridging two O6-atoms of guanine in a DNA duplex. When compared to the human AGT protein (hAGT), the S134P OGT variant displayed reduced activity towards O6-alkylation but a much broader substrate range for O4-alkylation damage reversal. The role of residue 134 in OGT is similar to its function in the human homolog, where Pro140 is crucial in conferring on hAGT the capability to repair large adducts at the O6-position of guanine. Finally, a method to generate a covalent conjugate between hAGT and a model nucleoside using a single-stranded oligonucleotide substrate is demonstrated. |
| topic |
Bioorganic molecules modified oligonucleotides DNA repair substrate specificity mutagenesis homology modeling DNA interstrand cross-link |
| url |
https://www.mdpi.com/1420-3049/22/11/1948 |
| work_keys_str_mv |
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