PDBsum entry 1jg0

Transferase

PDB id: 1jg0

Contents

Protein chains

263 a.a. *

Ligands

UMP ×2

DDT ×2

Waters ×398

* Residue conservation analysis

PDB id:

1jg0

Name:

Transferase

Title:

Crystal structure of escherichia coli thymidylate synthase complexed with 2'-deoxyuridine-5'-monophosphate and n,o-didansyl-l-tyrosine

Structure:

Thymidylate synthase. Chain: a, b. Engineered: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: esherichia coli. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PQS)

Resolution:

2.00Å R-factor: 0.212 R-free: 0.250

Authors:

T.A.Fritz,D.Tondi,J.S.Finer-Moore,M.P.Costi,R.M.Stroud

Key ref:

T.A.Fritz et al. (2001). Predicting and harnessing protein flexibility in the design of species-specific inhibitors of thymidylate synthase. Chem Biol, 8, 981-995. PubMed id: 11590022 DOI: 10.1016/S1074-5521(01)00067-9

Date:

22-Jun-01 Release date: 08-Feb-02

Protein chains

P0A884  (TYSY_ECOLI) -  Thymidylate synthase from Escherichia coli (strain K12)

Seq: 264 a.a.

Struc: 263 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.2.1.1.45 - thymidylate synthase.
• Pathway: Folate Coenzymes
• Reaction: dUMP + (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate = 7,8-dihydrofolate + dTMP

dUMP + (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate (Bound ligand (Het Group name = UMP) corresponds exactly) = 7,8-dihydrofolate + dTMP

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1016/S1074-5521(01)00067-9

Chem Biol 8:981-995 (2001)

PubMed id: 11590022

Predicting and harnessing protein flexibility in the design of species-specific inhibitors of thymidylate synthase.

T.A.Fritz, D.Tondi, J.S.Finer-Moore, M.P.Costi, R.M.Stroud.

ABSTRACT

BACKGROUND: Protein plasticity in response to ligand binding abrogates the notion of a rigid receptor site. Thus, computational docking alone misses important prospective drug design leads. Bacterial-specific inhibitors of an essential enzyme, thymidylate synthase (TS), were developed using a combination of computer-based screening followed by in-parallel synthetic elaboration and enzyme assay [Tondi et al. (1999) Chem. Biol. 6, 319-331]. Specificity was achieved through protein plasticity and despite the very high sequence conservation of the enzyme between species. RESULTS: The most potent of the inhibitors synthesized, N,O-didansyl-L-tyrosine (DDT), binds to Lactobacillus casei TS (LcTS) with 35-fold higher affinity and to Escherichia coli TS (EcTS) with 24-fold higher affinity than to human TS (hTS). To reveal the molecular basis for this specificity, we have determined the crystal structure of EcTS complexed with DDT and 2'-deoxyuridine-5'-monophosphate (dUMP). The 2.0 A structure shows that DDT binds to EcTS in a conformation not predicted by molecular docking studies and substantially differently than other TS inhibitors. Binding of DDT is accompanied by large rearrangements of the protein both near and distal to the enzyme's active site with movement of C alpha carbons up to 6 A relative to other ternary complexes. This protein plasticity results in novel interactions with DDT including the formation of hydrogen bonds and van der Waals interactions to residues conserved in bacterial TS but not hTS and which are hypothesized to account for DDT's specificity. The conformation DDT adopts when bound to EcTS explains the activity of several other LcTS inhibitors synthesized in-parallel with DDT suggesting that DDT binds to the two enzymes in similar orientations. CONCLUSIONS: Dramatic protein rearrangements involving both main and side chain atoms play an important role in the recognition of DDT by EcTS and highlight the importance of incorporating protein plasticity in drug design. The crystal structure of the EcTS/dUMP/DDT complex is a model system to develop more selective TS inhibitors aimed at pathogenic bacterial species. The crystal structure also suggests a general formula for identifying regions of TS and other enzymes that may be treated as flexible to aid in computational methods of drug discovery.

Selected figure(s)

Figure 5.
Fig. 5. DDT (N,O-didansyl-L-tyrosine) binding induces dramatic side chain and backbone plasticity. A: Divergent stereo view showing the plasticity of the side chains of active site residues between the EcTS/dUMP/DDT and EcTS/dUMP/ZD1694 structures. DDT and dUMP are shown in yellow, active site residues from the EcTS/dUMP/DDT structure are shown in atomic colors and active site residues from the EcTS/dUMP/ZD1694 structure are shown in gray. B: Divergent stereo view of aligned monomer backbones of the EcTS/dUMP/DDT (black and red) and EcTS/dUMP/ZD1694 (green) structures. The orientation of the structure is the same as that in A and DDT and dUMP are shown in yellow. Residues in red mapped onto the EcTS/dUMP/DDT backbone structure are approximately centrally located residues of protein segments which shift by > 1Å between the two structures. The location of the J-helix (J) is also indicated. C: Difference in Cα position between aligned structures of the EcTS/dUMP/DDT and EcTS/dUMP/ZD1694 complexes with the position of residues shown in B indicated.

Figure 6.
Fig. 6. Ligand-induced, main chain plastic accommodation of EcTS. Individual panels represent the positional difference between corresponding Cα carbons of the EcTS/dUMP/mTHF structure [12] aligned with (A) the EcTS/dUMP/ZD1694 [10], (B) the EcTS/dUMP/BW1843U89 [11], (C) the EcTS/dUMP/DDT or (D) the apo EcTS structure [17]. Only one monomer of each of the aligned structures is shown. D represents the structural changes known as segmental accommodation that take place upon cofactor binding to TS.

The above figures are reprinted by permission from Cell Press: Chem Biol (2001, 8, 981-995) copyright 2001.

Figures were selected by the author.

Author's comment

Protein plasticity in response to ligand binding abrogates the notion of a rigid receptor site. Hence computational docking alone misses important prospective drug design leads. Bacterial-specific inhibitors of an essential enzyme, thymidylate synthase (TS), were developed using a combination of computer-based screening followed by synthetic elaboration. The most potent of the inhibitors synthesized, N,O-didansyl-L-tyrosine (DDT), binds to Lactobacillus casei TS (LcTS) with 35-fold higher affinity and to bacterial TSs with ~30-fold higher affinity than to human TS (hTS). The 2.0Å structure shows that DDT binds to EcTS in a conformation not predicted by molecular docking. Binding of DDT is accompanied by large rearrangements of the protein both near and distal to the enzyme's active site with movement of C alpha carbons up to 6Å relative to other ternary complexes which account for DDT's specificity. Dramatic protein rearrangements involving both main and side chain atoms highlight the importance of incorporating protein plasticity in drug design. The crystal structure also suggests a general formula for identifying regions of proteins that may be treated as flexible to aid in computational methods of drug discovery.
Robert M. Stroud