Thymidylate Synthase

The Role of Protein Dynamics in Thymidylate Synthase Catalysis: Variants of Conserved 2'-Deoxyuridine 5'-Monophosphate (dUMP)-Binding Tyr-261. Zachary Newby, Tom T. Lee, Richard J. Morse, Yaoquan Liu, Lu Liu, Prasanna Venkatraman, Daniel V. Santi, Janet S. Finer-Moore, and Robert M. Stroud. (2006) Biochemistry Vol 45, 24.
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The crystal structure of dihydrofolate reductase-thymidylate synthase from Cryptosporidium hominis reveals a novel architecture for the bifunctional enzyme.
O'Neil, R. H., Lilien, R. H., Donald, B. R., Stroud, R.M. and Anderson, A.C. (2003) J Eukaryot Microbiol 50 Suppl p555-6.

Lessons and Conclusions from Dissecting the Mechanism of a Bi-substrate Enzyme: Thymidylate Synthase Mutagenesis, Function and Structure.
Finer-Moore, J.S., Santi, D.V., Stroud, R.M. (2003) Biochemistry 42, 248-256 (web released Dec 17th 2002 Bi 020599a)

Tryptophan 80 and leucine 143 are critical for the hydride transfer step of thymidylate synthase by controlling active site access.
Fritz TA, Liu L, Finer-Moore JS, Stroud RM. (2002). Biochemistry. 41, 7021-7025.
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The Only Active Mutant of Thymidylate Synthase D169, a Residue Far from the Site of Methyl Transfer, Demonstrates the Exquisite Nature of Catalysis in Enzymology.
Birdsall, D.L., Bencal, J., Santi, D.V., Stroud, R.M., and Finer-Moore-J. (2003).
Protein Engineering 16, 229-240.
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The structural roles of conserved Pro-196, Pro-197 and His-199 in the mechanism of thymidylate synthase.
Gonzalez-Pacanowska, D., Ruiz-Perez, L.M., Carreras-Gomez, M. A., Costi, M.P., Stroud, R.M., Finer-Moore, J.S. and Santi, D.V. (2003) Protein Engineering, 16, 229-240.
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Phylogenetic classification of protozoa based on the structure of the linker domain in the bifunctional enzyme, dihydrofolate reductase-thymidylate synthase.
O'Neil, R.H., Lilien, R.H., Donald, B.R., Stroud, R.M., and Anderson, A.C. (2003) J Biol Chem 278 52980-7.
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Structure-based studies on species-specific inhibition of thymidylate synthase.
Costi, M.P., Tondi, D., Rinaldi, M., Barlocco, D., Pecorari, P., Soragni, F., Venturelli, A., and Stroud, R.M. (2002) Biochem Biophys Acta 1587, 206-214
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Conformational Dynamics Along an Enzymatic Reaction Pathway: Thymidylate Synthase, 'the Movie.'
Robert M. Stroud, Janet S. Finer-Moore (2002) Biochemistry (web released Dec 17th 2002 Bi 020598i)
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Lessons and Conclusions from Dissecting the Mechanism of a Bi-substrate Enzyme: Thymidylate Synthase Mutagenesis, Function and Structure.
Finer-Moore, J.S., Santi, D.V., Stroud, R.M. (2002) Biochemistry (web released Dec 17th 2002 Bi 020599a)

Approaches to solving the rigid receptor problem by identifying a minimal set of flexible residues during ligand docking.
Anderson, A.C., O'Neil, R.H., Surti, T.S., Stroud, R.M. Chemistry & Biology 8 (200) 445-457.
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Enzyme-catalyzed therapeutic agent (ECTA) design: activation of the antitumor ECTA compound NB1011 by thymidylate synthase.
Lackey, D., Groziak, M., Sergeeva, M., Beryt, M., Boyer, C., Stroud, R.M., Sayre, P., Park, J., Johnston, P., Slamon, D., Shepard, M., Pegram, M. (2001) Biochemical Pharmacology 61: 179-189.
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Multi-targeted antifolates aimed at avoiding drug resistance form covalent closed inhibitory complexes with human and Escherichia coli thymidylate synthase.
Sayre, P.H., Finer-Moore, J.S., Fritz, T.A., Biermann, D., Gates, S.B., MacKellar, W.C., Patel, V.F., Stroud, R.M. (2001). Journal of Molecular Biology 313 4 813-829
Review abstract
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Predicting and harnessing protein flexibility in the design of species-specific inhibitors of thymidylate synthase.
Fritz, T.A., Tondi, D., Finer-Moore, J.S., Costi, M.P., and Stroud, R.M. (2001). Chemistry & Biology 8 No. 10 981-995.
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Effects of subunit occupancy on partitioning of an intermediate in thymidylate synthase mutants. Variath, P., Yaoquan Liu, Lee, T.T., Stroud, R.M., and Santi, D.V. (2000). Biochemistry 39, no. 10, 2429-2435.
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Replacement set mutagenesis of the four phosphate-binding arginine residues of thymidylate synthase.
Kawase, S., Cho, S.W., Rozelle, J., Stroud, R.M., Finer-Moore, J., and Santi, D. (2000). Protein Engineering. 13 8. 557-563.
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Site-directed ligand discovery.
Erlanson, D., Braisted, A., Raphael, D., Randal, M., Stroud, R.M., Gordon, E.M., Wells, J. (2000) PNAS 97: 17 9347-9372
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The crystal structure of thymidylate synthase from Pneumocystis carinii reveals a fungal insert important for drug design.
Anderson, A.C., Perry, K.M., Freymann, D.M., and Stroud, R.M. (2000). Journal of Molecular Biology 297(3): 645-657.
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Energetic contributions of four arginines to phosphate-binding in thymidylate synthase are more than additive and depend on optimization of effective charge balance.
Morse, R.J., Kawase, S., Santi, D.V., Finer-Moore, J., and Stroud, R. (2000). Biochemistry 39: 1011-1020.
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Conserved phosphate-binding arginines in thymidylate synthase: Mutations of Arg 178, but not Arg 179 or Arg 23, destabilize the phosphate binding site and impair dump binding.
Finer-Moore, JS; Fauman, EB; Morse, RJ; Santi, DV; Stroud, RM.(1999) Biochemistry
PDB Accession Nos. 1BO7, 1BO8, 1BPO, 1BP6 and 1BPJ
View PDB structure 1BO7, 1BO8, 1BP0, 1BP6, 1BPJ

The structural mechanism for half-the-sites reactivity in an enzyme, thymidylate synthase, involves a relay of changes between subunits.
Anderson, AC; O'Neil, RH; Delano, WL; Stroud, RM. (1999) Biochemistry 38, 13829-13836
PDB Accession No. 1CI7
Review abstract
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Phthalein derivatives as a new tool for selectivity in thymidylate synthase inhibition.
Costi, MP; Rinaldi, M; Tondi, D; Pecorari, P; Barlocco, D; Ghelli, S; Stroud, RM; Santi, DV; Stout, TJ; Musiu, C; Marangiu, EM; Pani, A; Congiu, D; Loi, GA; and La Colla, P. (1999) Journal of Medicinal Chemistry 42, 2112-2124.
Review abstract
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Structure-based design of inhibitors specific for bacterial thymidylate synthase.
Stout, TJ; Tondi, D; Rinaldi, M; Barlocco, D; Pecorari, P; Santi, DV; Kuntz, ID; Stroud, RM; Shoichet, BK; and Costi, MP. (1999)
Biochemistry 38, 1607-1617
PDB Accession Nos: 1TSL (LcTS with compound 4), 1TSM (LcTS with compound 5).
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View PDB structure 1TSL, 1TSM

Discovery of thymidylate synthase inhibitors using flexible docking and an empirically tuned scoring function.
Welch, W; Sage, CR; Stout, TJ; Klein, T; Ruppert, J; Stroud, RM; Jain, A. (1998)
PDB Accession No. 1AXW
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D221 in thymidylate synthase controls conformation change, and thereby opening of the imidazolidine.
Sage, CR; Michelitsch, MD; Stout, TJ; Biermann, D; Nissen, R; Finer-Moore, J; and Stroud, RM. (1998)
Biochemistry 37, 13893-13901.
PDB Accession Nos: 1BJG (D169N with FdUMP , CH2H4folate); 1DNA (D169N with dUMP, CB3717).
View PDB structure 1BJG, 1DNA
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Crystal structures of a unique thermal-stable thymidylate synthase from Bacillus subtilis.
Stout, TJ; Schellenberger, U; Santi, DV; and Stroud, RM. (1998) Biochemistry 37, 14736-14747.
PDB Accession Numbers:1BKP for bsTS-I, 1BKO for bsTS-II, 1BSP for bsTS-III, and 1BSF for bsTS-IV.
View PDB structure 1BKP, 1BKO, 1BSP, 1BSF
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The additivity of substrate fragments in enzyme-ligand binding.
Stout, TJ; Sage, CR; and Stroud, RM. (1998) Structure 6, 839-848.
PDB Accession Numbers:1TJS for apo-TS, 1BID for binaryTS + dUMP, 1BDU for binaryTS + dUrd + PO4,sup2; -, 1AOB for binaryTS + ddUrd + PO4,sup2; -, 1KCE for ternary TS + CB3717 + dUMP, 1TDU for ternary TS + CB3717 + dUrd + PO4,sup2; -, 1DDU for ternary TS + CB3717 + ddUrd + PO4,sup2; -, 1AN5 for ternary TS + CB3717 + PO4,sup2; -.
View PDB structure 1TJS, 1BID, 1BDU, 1AOB, 1KCE, 1TDU, 1DDU, 1AN5

Inactivity of N229A thymidylate synthase due to water mediated effects: Isolating a late stage in methyl transfer.
Reyes, CL; Sage, CR; Rutenber, EE; Nissen, R; Stroud, RM; and Finer-Moore JS. (1998). Journal of Molecular Biology 284, 699-712.
PDB Accession Nos:1BQ1 for ternary N229(177)A, 1BQ2 for binary N229(177)A.
View PDB structure 1BQ1, 1BQ2
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The separate effects of E60Q in Lactobacillus casei thymidylate synthase delineate between mechanisms for formation of intermediates in catalysis.
Birdsall, DL; Huang, W; Santi, DV; Stroud, RM; and Finer-Moore, J. (1998)
Protein Engineering 11, 171-183.
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Contributions of orientation and hydrogen bonding to catalysis in Asn229 mutants of thymidylate synthase.
Finer-Moore, JS; Liu, L; Birdsall, DL; Brem, R; Apfeld, J; Santi, DV; and Stroud, RM. (1998)
Journal of Molecular Biology 276, 113-129.
PDB Accession Nos: 1JMF, 1JMG, 1JMH, 1JMI, 1TVU, 1TVV , 1TVW.
View PDB structure 1JMF, 1JMG, 1JMH, 1JMI, 1TVU, 1TVV, 1TVW
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A novel dCMP methylase by engineering thymidylate synthase.
Agarwalla, S; LaPorte, S; Liu, L; Finer-Moore, J; Stroud, RM; and Santi, DV. (1997)
Biochemistry 36, 15909-15917.
Review abstract
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An essential role for water in an enzyme reaction mechanism: The crystal structure of the thymidylate synthase mutant E58Q.
Sage, CR; Rutenber, EE; Stout, TJ; and Stroud, RM. (1996)
Biochemistry 35, 16270-16281.
PDB Accession Nos. 1KCE (WT-TS), 1ZPR [E60(58)Q]
View PDB structure 1KCE, 1ZPR
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Binding of the anticancer drug ZD1694 to E. coli thymidylate synthase: assessing specificity and affinity.
Rutenber, EE; and Stroud, RM. (1996)
Structure 4, 1317-1324.
PDB Accession No. 2KCE
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Balancing ATP in the cell.
Stroud, RM. (1996)
Nature Structural Biology 3, 567-569
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Partitioning roles of side chains in affinity, orientation, and catalysis with structures for mutant complexes: asparagine-229 in thymidylate synthase.
Finer-Moore, JS; Liu, L; Schafmeister, CE; Birdsall, DL; Mau, T; Santi, DV; and Stroud, RM. (1996)
Biochemistry 35, 5125-5136.
PDB Accession Nos: 1NJA, 1NJB, 1NJC, 1NJD , 1NJE
View PDB structure 1NJA, 1NJB, 1NJC, 1NJD, 1NJE
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Asparagine 229 mutants of thymidylate synthase catalyze the methylation of 3-methyl-2'-deoxyuridine 5'-monophosphate.
Costi, PM; Liu, L; Finer-Moore, JS; Stroud, RM and Santi, DV. (1996)
Biochemistry 35, 3944-3949.
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Contribution of a salt bridge to binding affinity and dUMP orientation to catalytic rate: mutation of a substrate-binding arginine in thymidylate synthase.
Finer-Moore, JS; Fauman, EB; Morse, RJ; Santi, DV and Stroud, RM. (1996)
Protein Engineering 9, 69-75.
PDB Accession Nos: 1TSV, 1TSW, 1TSX, 1TSY, 1TSZ
View PDB structure 1TSV, 1TSW, 1TSX, 1TSY, 1TSZ
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Entropy in Bi-substrate enzymes - proposed role of an alternate site in chaperoning substrate into, and products out of, thymidylate synthase.
Birdsall, DL; Finer-Moore, J; and Stroud, RM. (1996)
Journal of Molecular Biology 255, 522-535.
PDB Accession Nos: 1VZA, 1VZB, 1VZC, 1VZD , 1VZE
View PDB structure 1VZA, 1VZB, 1VZC, 1VZD, 1VZE
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The complex of the anti-cancer therapeutic, BW1843U89, with thymidylate synthase at 2.0 Å resolution: Implications for a new mode of inhibition.
Stout, TJ; and Stroud, RM. (1996)
Structure 4, 67-77.
PDB Accession No: 1SYN
Review abstract
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Crystal structure of human thymidylate synthase: A structural mechanism for guiding substrates into the active site.
Schiffer, CA; Clifton, IJ; Davisson, VJ; Santi, DV; and Stroud, RM (1995).
Biochemistry 34, 16279-16287.
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Crystal structure of thymidylate synthase from T4 phage: component of a deoxynucleoside triphosphate-synthesizing complex.
Finer-Moore, JS; Maley, GF; Maley, F; Montfort, WR; and Stroud, RM. (1994)
Biochemistry 33, 15459-15468.
PDB Accession No. 1TIS
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An electrostatic highway.
Stroud, RM. (1994)
Nature Structural Biology 1, 131-134.
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Water-mediated substrate/product discrimination: The product complex of thymidylate synthase at 1.83 Å.
Fauman, EB; Rutenber, EE; Maley, G; Maley, F and Stroud, RM. (1994)
Biochemistry 33, 1502-1511.
PDB Accession No. 1TYS
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Refined structures of substrate-bound and phosphate-bound thymidylate synthase from Lactobacillus casei.
Finer-Moore, JS; Fauman, EB; Foster, PG; Perry, KM; Santi, DV; and Stroud, RM. (1993).
Journal of Molecular Biology 232, 1101-1116.
PDB Accession Nos: 1LCA, 1LCB, 1LCE, 2TDM (replaces 1TDM) , 1THY
View PDB structure 1LCA, 1LCB, 1LCE, 2TDM, 1THY
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Structures of thymidylate synthase with a C-terminal deletion: Role of the C-terminus in alignment of 2'-deoxyuridine 5'-monophosphate and 5,10-methylenetetrahydrofolate.
Perry, KM; Carreras, CW; Chang, LC; Santi, DV and Stroud, RM. (1993)
Biochemistry 32, 7116-7125.
PDB Accession Nos: 1TDA (V316Am inbound), 1TDB (V316Am·FdUMP), 1TDC (V316Am·dUMP), 2TDD (V316Am·FdUMP·5-HOCH2H4folate)
View PDB structure 1TDA, 1TDB, 1TDC, 2TDD
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Stereochemistry of a multistep/ bipartite methyl transfer reaction: thymidylate synthase.
Stroud, RM; and Finer-Moore, JS. (1993)
FASEB Journal 7, 671-677.
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Structure-based discovery of inhibitors of thymidylate synthase.
Shoichet, BK; Stroud, RM; Santi, DV; Kuntz, ID; and Perry, KM. (1993)
Science 259, 1445-1450.
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Cofactor triggers the conformational change in thymidylate synthase: implications for an ordered binding mechanism.
Kamb A; Finer-Moore JS; and Stroud RM. (1992)
Biochemistry 31, 12876-12884.

Structural basis for recognition of polyglutamyl folates by thymidylate synthase.
Kamb A; Finer-Moore J; Calvert AH; and Stroud RM. (1992)
Biochemistry 31, 9883-9890.
PDB Accession No. 2BBQ (replaces1BBQ referenced in Biochemistry 31, pp9883)
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Reversible dissociation and unfolding of the dimeric protein thymidylate synthase.
Perry, KM; Pookanjanatavip, M; Zhao, J; Santi, DV; Stroud, RM. (1992) Protein Science 1, 796-800.
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Crystallization of human thymidylate synthase.
Schiffer, CA; Davisson, VJ; Santi, DV and Stroud, RM. (1991) Journal of Molecular Biology 219, 161-163.
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Saturation site-directed mutagenesis of thymidylate.
Climie, S; Ruiz-Perez, L; Gonzalez-Pacanowska, D; Prapunwattana, P; Cho, SW; Stroud, R; and Santi, DV. (1990). Journal of Biological Chemistry 265, 18776-18779.
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Plastic adaptation toward mutations in proteins: structural comparison of thymidylate synthases.
Perry, KM; Fauman, EB; Finer-Moore, JS; Montfort, WR;
Maley, GF; Maley, F; and Stroud, RM. (1990)
Proteins 8, 315-333.
PDB Accession Nos. 3TMS , 4TMS
View PDB structure 3TMS, 4TMS
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Pairwise specificity and sequential binding in enzyme catalysis: thymidylate synthase.
Finer-Moore, JS; Montfort, WR; and Stroud, RM. (1990)
Biochemistry 29, 6977-6986.
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Structure, multiple site binding, and segmental accommodation in thymidylate synthase on binding dUMP and an anti-folate.
Montfort, WR; Perry, KM; Fauman, EB; Finer-Moore, JS; Maley, GF; Hardy, L; Maley, F; and Stroud, RM. (1990)
Biochemistry 29, 6964-6977.
PDB Accession No. 2TSC (replaces 1TSC , 1TMS referenced in Biochemistry 29, pp6964)
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Segmental accommodation: A novel conformational change induced upon ligand binding by thymidylate synthase.
Montfort, WR; Fauman, EB; Perry, KM; and Stroud, RM. (1990)
Current Research in Protein Chemistry Chapter 34, pp 367-382, Academic Press Inc.

Atomic structure of thymidylate synthase: Target for rational drug design.
Hardy, LW; Finer-Moore, JS; Montfort, WR; Jones, MO; Santi, DV; and Stroud, RM. (1987)
Science 235, 448-455.
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A pulsed diffusion technique for the growth of protein crystals for x-ray diffraction.
Koeppe, RE II; Stroud, RM; Pena, VA; and Santi, DV. (1975)
Journal of Molecular Biology 98, 155-160.
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