PDBsum entry 3fqf

Oxidoreductase

PDB id: 3fqf

Contents

Protein chain

157 a.a. *

Ligands

55V

NDP

Waters ×184

* Residue conservation analysis

PDB id:

3fqf

Name:

Oxidoreductase

Title:

Staphylococcus aureus f98y mutant dihydrofolate reductase complexed with NADPH and 2,4-diamino-5-[3-(3,4,5-trimethoxyphenyl)pent-1-ynyl]- 6-methylpyrimidine (ucp115a)

Structure:

Trimethoprim-sensitive dihydrofolate reductase. Chain: a. Engineered: yes. Mutation: yes

Source:

Staphylococcus aureus rf122. Organism_taxid: 273036. Strain: rf122 / et3-1. Gene: dfrb, sab1281c. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.77Å R-factor: 0.209 R-free: 0.245

Authors:

A.C.Anderson,K.M.Frey,J.Liu,M.N.Lombardo

Key ref:

K.M.Frey et al. (2009). Crystal structures of wild-type and mutant methicillin-resistant Staphylococcus aureus dihydrofolate reductase reveal an alternate conformation of NADPH that may be linked to trimethoprim resistance. J Mol Biol, 387, 1298-1308. PubMed id: 19249312 DOI: 10.1016/j.jmb.2009.02.045

Date:

07-Jan-09 Release date: 24-Mar-09

Protein chain

Q2YY41  (Q2YY41_STAAB) -

Enzyme reactions

• Enzyme class: E.C.1.5.1.3 - dihydrofolate reductase.
• Pathway: Folate Coenzymes
• Reaction: (6S)-5,6,7,8-tetrahydrofolate + NADP⁺ = 7,8-dihydrofolate + NADPH + H⁺

(6S)-5,6,7,8-tetrahydrofolate + NADP(+) (Bound ligand (Het Group name = NDP) corresponds exactly) = 7,8-dihydrofolate + NADPH + H(+)

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

reference

DOI no: 10.1016/j.jmb.2009.02.045

J Mol Biol 387:1298-1308 (2009)

PubMed id: 19249312

Crystal structures of wild-type and mutant methicillin-resistant Staphylococcus aureus dihydrofolate reductase reveal an alternate conformation of NADPH that may be linked to trimethoprim resistance.

K.M.Frey, J.Liu, M.N.Lombardo, D.B.Bolstad, D.L.Wright, A.C.Anderson.

ABSTRACT

Both hospital- and community-acquired Staphylococcus aureus infections have become major health concerns in terms of morbidity, suffering and cost. Trimethoprim-sulfamethoxazole (TMP-SMZ) is an alternative treatment for methicillin-resistant S. aureus (MRSA) infections. However, TMP-resistant strains have arisen with point mutations in dihydrofolate reductase (DHFR), the target for TMP. A single point mutation, F98Y, has been shown biochemically to confer the majority of this resistance to TMP. Using a structure-based approach, we have designed a series of novel propargyl-linked DHFR inhibitors that are active against several trimethoprim-resistant enzymes. We screened this series against wild-type and mutant (F98Y) S. aureus DHFR and found that several are active against both enzymes and specifically that the meta-biphenyl class of these inhibitors is the most potent. In order to understand the structural basis of this potency, we determined eight high-resolution crystal structures: four each of the wild-type and mutant DHFR enzymes bound to various propargyl-linked DHFR inhibitors. In addition to explaining the structure-activity relationships, several of the structures reveal a novel conformation for the cofactor, NADPH. In this new conformation that is predominantly associated with the mutant enzyme, the nicotinamide ring is displaced from its conserved location and three water molecules complete a network of hydrogen bonds between the nicotinamide ring and the protein. In this new position, NADPH has reduced interactions with the inhibitor. An equilibrium between the two conformations of NADPH, implied by their occupancies in the eight crystal structures, is influenced both by the ligand and the F98Y mutation. The mutation induced equilibrium between two NADPH-binding conformations may contribute to decrease TMP binding and thus may be responsible for TMP resistance.

Selected figure(s)

Figure 1.
Fig. 1. Stereoview images of the wild-type (teal) and Sa(F98Y) mutant (gold) enzymes bound to: a, compound 5 (wild-type light green, F98Y cyan); b, compound 8 (wild-type orange, F98Y lavender); c, compound 10 (wild-type purple, F98Y dark green); and d, compound 15 (wild-type yellow, F98Y gray). The standard conformation of NADPH is shown in magenta and the alternative conformation is shown in blue. Water molecules (shown in Fig. 3) are omitted for clarity.

Figure 2.
Fig. 2. Electron density at the active site for SaDHFR(F98Y):NADPH:5. Protein is shown with 2F[o]–F[c] density (1.5 σ, blue) and ligands are shown with omit F[o]–F[c] density (3.0 σ, grey). A full view of the density for the alternative conformation is shown at the right.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2009, 387, 1298-1308) copyright 2009.

Figures were selected by the author.