2drc

<StructureSection load='2drc' size='340' side='right' caption='[[2drc]], [[Resolution|resolution]] 1.90&Aring;' scene=''>

<table><tr><td colspan='2'>[[2drc]] is a 2 chain structure with sequence from [http://en.wikipedia.org/wiki/"bacillus_coli"_migula_1895 "bacillus coli" migula 1895]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2DRC OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=2DRC FirstGlance]. <br>

</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=CA:CALCIUM+ION'>CA</scene>, <scene name='pdbligand=CL:CHLORIDE+ION'>CL</scene>, <scene name='pdbligand=MTX:METHOTREXATE'>MTX</scene></td></tr>

<tr id='activity'><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/Dihydrofolate_reductase Dihydrofolate reductase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.5.1.3 1.5.1.3] </span></td></tr>

<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=2drc FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=2drc OCA], [http://pdbe.org/2drc PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=2drc RCSB], [http://www.ebi.ac.uk/pdbsum/2drc PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=2drc ProSAT]</span></td></tr>

[[http://www.uniprot.org/uniprot/DYR_ECOLI DYR_ECOLI]] Key enzyme in folate metabolism. Catalyzes an essential reaction for de novo glycine and purine synthesis, and for DNA precursor synthesis.

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== Publication Abstract from PubMed ==

We have applied site-directed mutagenesis methods to change the conserved tryptophan-22 in the substrate binding site of Escherichia coli dihydrofolate reductase to phenylalanine (W22F) and histidine (W22H). The crystal structure of the W22F mutant in a binary complex with the inhibitor methotrexate has been refined at 1.9-A resolution. The W22F difference Fourier map and least-squares refinement show that structural effects of the mutation are confined to the immediate vicinity of position 22 and include an unanticipated 0.4-A movement of the methionine-20 side chain. A conserved bound water-403, suspected to play a role in the protonation of substrate DHF, has not been displaced by the mutation despite the loss of a hydrogen bond with tryptophan-22. Steady-state kinetics, stopped-flow kinetics, and primary isotope effects indicate that both mutations increase the rate of product tetrahydrofolate release, the rate-limiting step in the case of the wild-type enzyme, while slowing the rate of hydride transfer to the point where it now becomes at least partially rate determining. Steady-state kinetics show that below pH 6.8, kcat is elevated by up to 5-fold in the W22F mutant as compared with the wild-type enzyme, although kcat/Km(dihydrofolate) is lower throughout the observed pH range. For the W22H mutant, both kcat and kcat/Km(dihydrofolate) are substantially lower than the corresponding wild-type values. While both mutations weaken dihydrofolate binding, cofactor NADPH binding is not significantly altered. Fitting of the kinetic pH profiles to a general protonation scheme suggests that the proton affinity of dihydrofolate may be enhanced upon binding to the enzyme. We suggest that the function of tryptophan-22 may be to properly position the side chain of methionine-20 with respect to N5 of the substrate dihydrofolate.

 

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== References ==

[[Category: Bacillus coli migula 1895]]

[[Category: Dihydrofolate reductase]]

[[Category: Brown, K A]]

[[Category: Kraut, J]]

[[Category: Oxidoreductase]]