Alpha-carboxy nucleoside phosphonates as universal nucleoside triphosphate mimics.
Polymerases have a structurally highly conserved negatively charged amino acid motif that is strictly required for Mg(2+) cation-dependent catalytic incorporation of (d)NTP nucleotides into nucleic acids. Based on these characteristics, a nucleoside monophosphonate scaffold, α-carboxy nucleoside phosphonate (α-CNP), was designed that is recognized by a variety of polymerases. Kinetic, biochemical, and crystallographic studies with HIV-1 reverse transcriptase revealed that α-CNPs mimic the dNTP binding through a carboxylate oxygen, two phosphonate oxygens, and base-pairing with the template. In particular, the carboxyl oxygen of the α-CNP acts as the potential equivalent of the α-phosphate oxygen of dNTPs and two oxygens of the phosphonate group of the α-CNP chelate Mg(2+), mimicking the chelation by the β- and γ-phosphate oxygens of dNTPs. α-CNPs (i) do not require metabolic activation (phosphorylation), (ii) bind directly to the substrate-binding site, (iii) chelate one of the two active site Mg(2+) ions, and (iv) reversibly inhibit the polymerase catalytic activity without being incorporated into nucleic acids. In addition, α-CNPs were also found to selectively interact with regulatory (i.e., allosteric) Mg(2+)-dNTP-binding sites of nucleos(t)ide-metabolizing enzymes susceptible to metabolic regulation. α-CNPs represent an entirely novel and broad technological platform for the development of specific substrate active- or regulatory-site inhibitors with therapeutic potential.
Rega Institute for Medical Research and jan.balzarini@rega.kuleuven.be.
Jan
J
Balzarini
Center for Advanced Biotechnology and Medicine and Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854-8020;
Kalyan
K
Das
Department of Biochemistry, McGill University, Montreal, QC H3G 1Y6, Canada;
Jean A
JA
Bernatchez
Center for Advanced Biotechnology and Medicine and Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854-8020;
Sergio E
SE
Martinez
Department of Microbiology and Immunology, McGill University, Montreal, QC H3A 2B4, Canada; Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, AB T6G 2E1, Canada;
Marianne
M
Ngure
Department of Chemistry and School of Pharmacy, Analytical and Biological Chemistry Research Facility, Synthesis and Solid State Pharmaceutical Centre, University College Cork, Cork, Ireland; and.
Sarah
S
Keane
Department of Chemistry and School of Pharmacy, Analytical and Biological Chemistry Research Facility, Synthesis and Solid State Pharmaceutical Centre, University College Cork, Cork, Ireland; and.
Alan
A
Ford
Department of Chemistry and School of Pharmacy, Analytical and Biological Chemistry Research Facility, Synthesis and Solid State Pharmaceutical Centre, University College Cork, Cork, Ireland; and.
Nuala
N
Maguire
Department of Chemistry and School of Pharmacy, Analytical and Biological Chemistry Research Facility, Synthesis and Solid State Pharmaceutical Centre, University College Cork, Cork, Ireland; and.
Niki
N
Mullins
Department of Chemistry, KU Leuven, B-3000 Leuven, Belgium;
Jubi
J
John
Department of Chemistry, KU Leuven, B-3000 Leuven, Belgium;
Youngju
Y
Kim
Department of Chemistry, KU Leuven, B-3000 Leuven, Belgium;
Wim
W
Dehaen
Rega Institute for Medical Research and.
Johan
J
Vande Voorde
Rega Institute for Medical Research and.
Sandra
S
Liekens
Rega Institute for Medical Research and.
Lieve
L
Naesens
Department of Biochemistry, McGill University, Montreal, QC H3G 1Y6, Canada; Department of Microbiology and Immunology, McGill University, Montreal, QC H3A 2B4, Canada; Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, AB T6G 2E1, Canada; Department of Medicine, Division of Experimental Medicine, McGill University, Montreal, QC H3A 1A3, Canada.
Matthias
M
Götte
Department of Chemistry and School of Pharmacy, Analytical and Biological Chemistry Research Facility, Synthesis and Solid State Pharmaceutical Centre, University College Cork, Cork, Ireland; and.
Anita R
AR
Maguire
Center for Advanced Biotechnology and Medicine and Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854-8020;
Eddy
E
Arnold
D000494:Allosteric Regulation / Q000187:drug effects; D001483:Base Sequence; D055162:Biocatalysis / Q000187:drug effects; D002457:Cell Extracts; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D024882:Drug Resistance, Viral / Q000187:drug effects; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009705:Nucleosides / Q000737:chemistry / Q000494:pharmacology*; D009711:Nucleotides / Q000737:chemistry / Q000494:pharmacology*; D063065:Organophosphonates / Q000737:chemistry / Q000494:pharmacology*; D018894:Reverse Transcriptase Inhibitors / Q000737:chemistry / Q000494:pharmacology; D013237:Stereoisomerism
D002457:Cell Extracts; D009705:Nucleosides; D009711:Nucleotides; D063065:Organophosphonates; D018894:Reverse Transcriptase Inhibitors; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase; D004259:DNA-Directed DNA Polymerase
(deoxy)nucleoside triphosphate mimic; HIV reverse transcriptase; allosteric inhibition; alpha-carboxy nucleoside phosphonate; herpes virus DNA polymerase