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| | ==DosS GAFA Domain Reduced Nitric Oxide Bound Crystal Structure== | | ==DosS GAFA Domain Reduced Nitric Oxide Bound Crystal Structure== |
| - | <StructureSection load='4yof' size='340' side='right' caption='[[4yof]], [[Resolution|resolution]] 1.90Å' scene=''> | + | <StructureSection load='4yof' size='340' side='right'caption='[[4yof]], [[Resolution|resolution]] 1.90Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| | <table><tr><td colspan='2'>[[4yof]] is a 2 chain structure with sequence from [http://en.wikipedia.org/wiki/Mycto Mycto]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4YOF OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4YOF FirstGlance]. <br> | | <table><tr><td colspan='2'>[[4yof]] is a 2 chain structure with sequence from [http://en.wikipedia.org/wiki/Mycto Mycto]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4YOF OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4YOF FirstGlance]. <br> |
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| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
| - | Mycobacterium tuberculosis (Mtb) DosS is critical for the induction of Mtb dormancy genes in response to nitric oxide (NO), carbon monoxide (CO) or hypoxia. These environmental stimuli, which are sensed by the DosS heme group, result in autophosphorylation of a DosS His residue, followed by phosphotransfer to an Asp residue of the response regulator DosR. To clarify the mechanism of gaseous ligand recognition and signaling, we investigated the hydrogen-bonding interactions of the iron-bound CO and NO ligands by site-directed mutagenesis of Glu87 and His89. Autophosphorylation assays and molecular dynamics simulations suggest that Glu87 has an important role in ligand recognition, while His89 is essential for signal transduction to the kinase domain, a process for which Arg204 is important. Mutation of Glu87 to Ala or Gly rendered the protein constitutively active as a kinase, but with lower autophosphorylation activity than the wild-type in the Fe(II) and the Fe(II)-CO states, whereas the E87D mutant had little kinase activity except for the Fe(II)-NO complex. The H89R mutant exhibited attenuated autophosphorylation activity, while the H89A and R204A mutants were inactive as kinases, emphasizing the importance of these residues in communication to the kinase core. Resonance Raman of the wild-type and H89A mutant indicate the mutation does not alter the heme coordination number, spin state, or porphyrin deformation state, but suggests that interdomain interactions are disrupted by the mutation. Overall, these results confirm the importance of the distal hydrogenbonding network in ligand recognition and communication to the kinase domain, and reveal the sensitivity of the system to subtle differences in the binding of gaseous ligands.
| + | Defining the conformational states of cytochrome P450 active sites is critical for the design of agents that minimize drug-drug interactions, the development of isoform-specific P450 inhibitors, and the engineering of novel oxidative catalysts. We used two-dimensional (1)H,(15)N HSQC chemical shift perturbation mapping of (15)N-labeled Phe residues and x-ray crystallography to examine the ligand-dependent conformational dynamics of CYP119. Active site Phe residues were most affected by the binding of azole inhibitors and fatty acid substrates, in agreement with active site localization of the conformational changes. This was supported by crystallography, which revealed movement of the F-G loop with various azoles. Nevertheless, the NMR chemical shift perturbations caused by azoles and substrates were distinguishable. The absence of significant chemical shift perturbations with several azoles revealed binding of ligands to an open conformation similar to that of the ligand-free state. In contrast, 4-phenylimidazole caused pronounced NMR changes involving Phe-87, Phe-144, and Phe-153 that support the closed conformation found in the crystal structure. The same closed conformation is observed by NMR and crystallography with a para-fluoro substituent on the 4-phenylimidazole, but a para-chloro or bromo substituent engendered a second closed conformation. An open conformation is thus favored in solution with many azole ligands, but para-substituted phenylimidazoles give rise to two closed conformations that depend on the size of the para-substituent. The results suggest that ligands selectively stabilize discrete cytochrome P450 conformational states. |
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| - | Distal Hydrogen-Bonding Interactions in Ligand Sensing and Signaling by Mycobacterium tuberculosis DosS.,Basudhar D, Madrona Y, Yukl ET, Sivaramakrishnan S, Nishida CR, Moenne-Loccoz P, Ortiz de Montellano PR J Biol Chem. 2016 May 27. pii: jbc.M116.724815. PMID:27235395<ref>PMID:27235395</ref>
| + | Analysis of cytochrome P450 CYP119 ligand-dependent conformational dynamics by two-dimensional NMR and X-ray crystallography.,Basudhar D, Madrona Y, Kandel S, Lampe JN, Nishida CR, de Montellano PR J Biol Chem. 2015 Apr 17;290(16):10000-17. doi: 10.1074/jbc.M114.627935. Epub, 2015 Feb 10. PMID:25670859<ref>PMID:25670859</ref> |
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| | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> |
| | </div> | | </div> |
| | <div class="pdbe-citations 4yof" style="background-color:#fffaf0;"></div> | | <div class="pdbe-citations 4yof" style="background-color:#fffaf0;"></div> |
| | + | |
| | + | ==See Also== |
| | + | *[[Response regulator 3D structure|Response regulator 3D structure]] |
| | == References == | | == References == |
| | <references/> | | <references/> |
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| | </StructureSection> | | </StructureSection> |
| | [[Category: Histidine kinase]] | | [[Category: Histidine kinase]] |
| | + | [[Category: Large Structures]] |
| | [[Category: Mycto]] | | [[Category: Mycto]] |
| | [[Category: Madrona, Y]] | | [[Category: Madrona, Y]] |
| Structural highlights
4yof is a 2 chain structure with sequence from Mycto. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
| | Ligands: | , |
| Related: | 4ynr, 2w3d |
| Gene: | devS, dosS, MT3218 (MYCTO) |
| Activity: | Histidine kinase, with EC number 2.7.13.3 |
| Resources: | FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT |
Function
[DEVS_MYCTO] Member of the two-component regulatory system DevR/DevS (DosR/DosS) involved in onset of the dormancy response. May act as a redox sensor (rather than a direct hypoxia sensor); the normal (aerobic growth) state is the Fe(3+) form, while the reduced (anaerobic growth) Fe(2+) form is probably active for phosphate transfer. It is probably reduced by flavin nucleotides such as FMN and FAD. May be the primary sensor for CO. Donates a phosphate group to DevR (DosR) (By similarity).
Publication Abstract from PubMed
Defining the conformational states of cytochrome P450 active sites is critical for the design of agents that minimize drug-drug interactions, the development of isoform-specific P450 inhibitors, and the engineering of novel oxidative catalysts. We used two-dimensional (1)H,(15)N HSQC chemical shift perturbation mapping of (15)N-labeled Phe residues and x-ray crystallography to examine the ligand-dependent conformational dynamics of CYP119. Active site Phe residues were most affected by the binding of azole inhibitors and fatty acid substrates, in agreement with active site localization of the conformational changes. This was supported by crystallography, which revealed movement of the F-G loop with various azoles. Nevertheless, the NMR chemical shift perturbations caused by azoles and substrates were distinguishable. The absence of significant chemical shift perturbations with several azoles revealed binding of ligands to an open conformation similar to that of the ligand-free state. In contrast, 4-phenylimidazole caused pronounced NMR changes involving Phe-87, Phe-144, and Phe-153 that support the closed conformation found in the crystal structure. The same closed conformation is observed by NMR and crystallography with a para-fluoro substituent on the 4-phenylimidazole, but a para-chloro or bromo substituent engendered a second closed conformation. An open conformation is thus favored in solution with many azole ligands, but para-substituted phenylimidazoles give rise to two closed conformations that depend on the size of the para-substituent. The results suggest that ligands selectively stabilize discrete cytochrome P450 conformational states.
Analysis of cytochrome P450 CYP119 ligand-dependent conformational dynamics by two-dimensional NMR and X-ray crystallography.,Basudhar D, Madrona Y, Kandel S, Lampe JN, Nishida CR, de Montellano PR J Biol Chem. 2015 Apr 17;290(16):10000-17. doi: 10.1074/jbc.M114.627935. Epub, 2015 Feb 10. PMID:25670859[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Basudhar D, Madrona Y, Kandel S, Lampe JN, Nishida CR, de Montellano PR. Analysis of cytochrome P450 CYP119 ligand-dependent conformational dynamics by two-dimensional NMR and X-ray crystallography. J Biol Chem. 2015 Apr 17;290(16):10000-17. doi: 10.1074/jbc.M114.627935. Epub, 2015 Feb 10. PMID:25670859 doi:http://dx.doi.org/10.1074/jbc.M114.627935
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