Effects of Acyclovir, Foscarnet, and Ribonucleotides on Herpes Simplex Virus-1 DNA Polymerase: Mechanistic Insights and a Novel Mechanism for Preventing Stable Incorporation of Ribonucleotides into DNA

Abstract

We examined the impact of two clinically approved anti-herpes drugs, acyclovir and Forscarnet (phosphonoformate) on the exonuclease activity of the herpes simplex virus-1 DNA polymerase, UL30. Acyclovir triphosphate and Foscarnet, along with the closely related phosphonoacetic acid, did not affect exonuclease activity on single-stranded DNA. Furthermore, blocking the polymerase active site due to either binding of Foscarnet or phosphonoacetic acid to the E-DNA complex or polymerization of acyclovir onto the DNA also had minimal effect on exonuclease activity. The inability of the exonuclease to excise acyclovir from the primer 3′-terminus results from the altered sugar structure directly impeding phosphodiester bond hydrolysis as opposed to inhibiting binding, unwinding of the DNA by the exonuclease or transfer of the DNA from the polymerase to the exonuclease. Removing the 3′-hydroxyl or the 2′-carbon from the nucleotide at the 3′-terminus of the primer strongly inhibited exonuclease activity, although addition of a 2′-hydroxyl did not affect exonuclease activity. The biological consequences of these results are two-fold. First, the ability of acyclovir and Foscarnet to block dNTP polymerization without impacting exonuclease activity raises the possibility that their effects on herpes replication may involve both direct inhibition of dNTP polymerization along with exonuclease mediated destruction of herpes DNA. Second, the ability of the exonuclease to rapidly remove a ribonucleotide at the primer 3´-terminus in combination with the polymerase not efficiently adding dNTPs onto this primer provides a novel mechanism by which the herpes replication machinery can prevent incorporation of ribonucleotides into newly synthesized DNA.

Acute renal injury induced by valacyclovir hydrochloride: A case report

“…It directly causes cell membrane injury, alters the membrane permeability and ion transportation. In addition, it destroys cytoplasm mitochondria, inhibits enzymatic activity and protein synthesis (21), promotes calcium internal flow and leads to cytoskeleton structural damage and epithelial cell necrosis. Furthermore, it is able to produce oxygen free radicals.…”

“…Fourthly, it may cause thrombotic microangiopathy (23). Fifthly, the physiological nature of the kidney is highly susceptible to drug-induced nephropathy (21). The abundant renal capillary network and rich blood allow for high concentrations of drugs passing through a large contacted area.…”

Distribution and effects of amino acid changes in drug-resistant α and β herpesviruses DNA polymerase

“…ACV triphosphate (ACV-TP) competitively inhibits dGTP binding and is incorporated into DNA by HSV-1/-2 and VZV DNApol to act as a chain terminator (Figure 6A). ACV incorporated at the 3′ extremity of the elongated DNA is also considered a suicide inhibitor because the viral DNApol cannot bypass its presence through the 3′-5′exonuclease activity (43,44). Although GCV triphosphate (GCV-TP) is structurally related to ACV-TP, its mode of action is slightly different (Figure 6A).…”

Different Divalent Cations Alter the Kinetics and Fidelity of DNA Polymerases

“…All DNA polymerases (DNA pols) 2 require Mg 2ϩ or Mn 2ϩ for primer extension and for excision of incorrectly incorporated dNTPs via intrinsic 3Ј35Ј exonuclease activity (1)(2)(3)(4)(5). A two-metal-ion mechanism is used by all DNA pols to catalyze nucleotide addition to a growing primer strand (6).…”