2inq
From Proteopedia
(New page: 200px<br /><applet load="2inq" size="450" color="white" frame="true" align="right" spinBox="true" caption="2inq, resolution 2.20Å" /> '''Neutron Crystal Stru...) |
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| - | [[Image:2inq.gif|left|200px]]<br /><applet load="2inq" size=" | + | [[Image:2inq.gif|left|200px]]<br /><applet load="2inq" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="2inq, resolution 2.20Å" /> | caption="2inq, resolution 2.20Å" /> | ||
'''Neutron Crystal Structure of Escherichia coli Dihydrofolate Reductase Bound to the Anti-cancer drug, Methotrexate'''<br /> | '''Neutron Crystal Structure of Escherichia coli Dihydrofolate Reductase Bound to the Anti-cancer drug, Methotrexate'''<br /> | ||
==Overview== | ==Overview== | ||
| - | Hydrogen atoms play a central role in many biochemical processes yet are | + | Hydrogen atoms play a central role in many biochemical processes yet are difficult to visualize by x-ray crystallography. Spallation neutron sources provide a new arena for protein crystallography with TOF measurements enhancing data collection efficiency and allowing hydrogen atoms to be located in smaller crystals of larger biological macromolecules. Here we report a 2.2-A resolution neutron structure of Escherichia coli dihydrofolate reductase (DHFR) in complex with methotrexate (MTX). Neutron data were collected on a 0.3-mm(3) D(2)O-soaked crystal at the Los Alamos Neutron Scattering Center. This study provides an example of using spallation neutrons to study protein dynamics, to identify protonation states directly from nuclear density maps, and to analyze solvent structure. Our structure reveals that the occluded loop conformation [monomer (mon.) A] of the DHFR.MTX complex undergoes greater H/D exchange compared with the closed-loop conformer (mon. B), partly because the Met-20 and beta(F-G) loops readily exchange in mon. A. The eight-stranded beta sheet of both DHFR molecules resists H/D exchange more than the helices and loops. However, the C-terminal strand, betaH, in mon. A is almost fully exchanged. Several D(2)Os form hydrogen bonds with exchanged amides. At the active site, the N1 atom of MTX is protonated and thus charged when bound to DHFR. Several D(2)Os are observed at hydrophobic surfaces, including two pockets near the MTX-binding site. A previously unidentified D(2)O hydrogen bonds with the catalytic D27 in mon. B, stabilizing its negative charge. |
==About this Structure== | ==About this Structure== | ||
| - | 2INQ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with MT1 as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [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] Full crystallographic information is available from [http:// | + | 2INQ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with <scene name='pdbligand=MT1:'>MT1</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [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] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2INQ OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Escherichia coli]] | [[Category: Escherichia coli]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Bennett, B | + | [[Category: Bennett, B C.]] |
[[Category: Coates, L.]] | [[Category: Coates, L.]] | ||
| - | [[Category: Dealwis, C | + | [[Category: Dealwis, C G.]] |
| - | [[Category: Langan, P | + | [[Category: Langan, P A.]] |
[[Category: Schoenborn, B.]] | [[Category: Schoenborn, B.]] | ||
[[Category: MT1]] | [[Category: MT1]] | ||
[[Category: neutron structure; deuterium exchange; pseudo-rossman fold; nucleotide binding domain; chemotherapy]] | [[Category: neutron structure; deuterium exchange; pseudo-rossman fold; nucleotide binding domain; chemotherapy]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 17:54:23 2008'' |
Revision as of 15:54, 21 February 2008
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Neutron Crystal Structure of Escherichia coli Dihydrofolate Reductase Bound to the Anti-cancer drug, Methotrexate
Overview
Hydrogen atoms play a central role in many biochemical processes yet are difficult to visualize by x-ray crystallography. Spallation neutron sources provide a new arena for protein crystallography with TOF measurements enhancing data collection efficiency and allowing hydrogen atoms to be located in smaller crystals of larger biological macromolecules. Here we report a 2.2-A resolution neutron structure of Escherichia coli dihydrofolate reductase (DHFR) in complex with methotrexate (MTX). Neutron data were collected on a 0.3-mm(3) D(2)O-soaked crystal at the Los Alamos Neutron Scattering Center. This study provides an example of using spallation neutrons to study protein dynamics, to identify protonation states directly from nuclear density maps, and to analyze solvent structure. Our structure reveals that the occluded loop conformation [monomer (mon.) A] of the DHFR.MTX complex undergoes greater H/D exchange compared with the closed-loop conformer (mon. B), partly because the Met-20 and beta(F-G) loops readily exchange in mon. A. The eight-stranded beta sheet of both DHFR molecules resists H/D exchange more than the helices and loops. However, the C-terminal strand, betaH, in mon. A is almost fully exchanged. Several D(2)Os form hydrogen bonds with exchanged amides. At the active site, the N1 atom of MTX is protonated and thus charged when bound to DHFR. Several D(2)Os are observed at hydrophobic surfaces, including two pockets near the MTX-binding site. A previously unidentified D(2)O hydrogen bonds with the catalytic D27 in mon. B, stabilizing its negative charge.
About this Structure
2INQ is a Single protein structure of sequence from Escherichia coli with as ligand. Active as Dihydrofolate reductase, with EC number 1.5.1.3 Full crystallographic information is available from OCA.
Reference
Neutron diffraction studies of Escherichia coli dihydrofolate reductase complexed with methotrexate., Bennett B, Langan P, Coates L, Mustyakimov M, Schoenborn B, Howell EE, Dealwis C, Proc Natl Acad Sci U S A. 2006 Dec 5;103(49):18493-8. Epub 2006 Nov 27. PMID:17130456
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