Daniel V Santi

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Thymidylate synthase (TS, EC catalyzes the reductive methylation of dUMP by CH2H4folate to produce dTMP and H2folate. Knowledge of the catalytic mechanism and structure of TS has increased substantially over recent years. Major advances were derived from crystal structures of TS bound to various ligands, the ability to overexpress TS in(More)
Single and multiple mutations at residues 16, 51, 59, 108, and 164 of Plasmodium falciparum dihydrofolate reductase (pfDHFR) have been linked to antifolate resistance in malaria. We prepared and characterized all seven of the pfDHFR mutants found in nature, as well as six mutants not observed in nature. Mutations involving residues 51, 59, 108, or 164(More)
DNA containing 5-azacytosine (azaC) has previously been shown to be a potent inhibitor of DNA-cytosine methyltransferases. In this report, we describe experiments which demonstrate that azaC-DNA forms a covalent complex with Hpa II methylase, a bacterial enzyme that methylates the internal C of C-C-G-G sequences. The complex does not undergo detectable(More)
The polyketides are a diverse group of natural products with great significance as human and veterinary pharmaceuticals. A significant barrier to the production of novel genetically engineered polyketides has been the lack of available heterologous expression systems for functional polyketide synthases (PKSs). Herein, we report the expression of an intact(More)
The complete nucleotide sequences of the Haemophilus influenzae and Mycoplasma genitalium genomes and the partially sequenced Escherichia coli chromosome were analyzed to identify open reading frames (ORFs) likely to encode RNA modifying enzymes. The protein sequences of known RNA modifying enzymes from three families--m5U methyltransferases, psi synthases(More)
To exploit the huge potential of whole-genome sequence information, the ability to efficiently synthesize long, accurate DNA sequences is becoming increasingly important. An approach proposed toward this end involves the synthesis of approximately 5-kb segments of DNA, followed by their assembly into longer sequences by conventional cloning methods [Smith,(More)
A computer modeling procedure for assessing the stereochemical suitability of pairs of residues in proteins as potential sites for introduction of cystine disulfide crosslinks has been developed. Residue pairs with C alpha-C alpha distances of less than or equal to 6.5 A and C beta-C beta distances of less than or equal to 4.5 A are chosen for geometrical(More)
A barrier to heterologous production of complex polyketides in Escherichia coli is the lack of (2S)-methylmalonyl-CoA, a common extender substrate for the biosynthesis of complex polyketides by modular polyketide synthases. One biosynthetic route to (2S)-methylmalonyl-CoA involves the sequential actions of two enzymes, methylmalonyl-CoA mutase and(More)
Kinetic and catalytic properties of the DNA (cytosine-5)-methyltransferase HhaI are described. With poly(dG-dC) as substrate, the reaction proceeds by an equilibrium (or processive) ordered Bi-Bi mechanism in which DNA binds to the enzyme first, followed by S-adenosylmethionine (AdoMet). After methyl transfer, S-adenosylhomocysteine (AdoHcy) dissociates(More)