1mto

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(New page: 200px<br /><applet load="1mto" size="450" color="white" frame="true" align="right" spinBox="true" caption="1mto, resolution 3.20&Aring;" /> '''X-ray Crystal struct...)
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[[Image:1mto.gif|left|200px]]<br /><applet load="1mto" size="450" color="white" frame="true" align="right" spinBox="true"
 
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caption="1mto, resolution 3.20&Aring;" />
 
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'''X-ray Crystal structure of a Phosphofructokinase mutant from Bacillus stearothermophilus bound with frutose-6-phosphate'''<br />
 
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==Overview==
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==Crystal structure of a Phosphofructokinase mutant from Bacillus stearothermophilus bound with fructose-6-phosphate==
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The biophysical properties of a tryptophan-shifted mutant of, phosphofructokinase from Bacillus stearothermophilus (BsPFK) have been, examined. The mutant, designated W179Y/Y164W, has kinetic and, thermodynamic properties similar to the wild-type enzyme. A 2-fold, decrease in kcat is observed, and the mutant displays a 3-fold smaller, K(0.5) for the substrate, fructose-6-phosphate (Fru-6-P), as compared to, the wild-type enzyme. The dissociation constant for the inhibitor, phospho(enol)pyruvate (PEP), increases 2-fold, and the coupling parameter, Q(ay), decreases 2-fold. This suggests that while the mutant displays a, slightly decreased affinity for PEP, PEP is still an effective inhibitor, once bound. The new position of the tryptophan in W179Y/Y164W is, approximately 6 A from the Fru-6-P portion of the active site. A 25%, decrease in fluorescence intensity is observed upon Fru-6-P binding, and, an 80% decrease in fluorescence intensity is observed with PEP binding. In, addition, the intrinsic fluorescence polarization increases from 0.327 +/-, 0.001 to 0.353 +/- 0.001 upon Fru-6-P binding, but decreases to 0.290 +/-, 0.001 when PEP binds. Most notably, the presence of PEP induces, dissociation of the tetramer. Dissociation of the tetramer into dimers, occurs along the active site interface and can be monitored by the loss in, activity or the loss in tryptophan fluorescence that is observed when the, enzyme is titrated with PEP. Activity can be protected or recovered by, incubating the enzyme with Fru-6-P. Recovery of activity is enzyme, concentration dependent, and the rate constant for association is 6.2 +/-, 0.3 M(-1) x s(-1). Ultracentrifugation experiments revealed that in the, absence of PEP the mutant enzyme exists in an equilibrium between the, dimer and tetramer forms with a dissociation constant of 11.8 +/- 0.5, microM, while in the presence of PEP the enzyme exists in equilibrium, between the dimer and monomer forms with a dissociation constant of 7.5, +/- 0.02 microM. A 3.1 A crystal structure of the mutant enzyme suggests, that the amino acid substitutions have not dramatically altered the, tertiary structure of the enzyme. While it is clear that wild-type BsPFK, exists as a tetramer under these same conditions, these results suggest, that quaternary structural changes probably play an important role in, allosteric communication.
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<StructureSection load='1mto' size='340' side='right'caption='[[1mto]], [[Resolution|resolution]] 3.20&Aring;' scene=''>
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== Structural highlights ==
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<table><tr><td colspan='2'>[[1mto]] is a 8 chain structure with sequence from [https://en.wikipedia.org/wiki/Geobacillus_stearothermophilus Geobacillus stearothermophilus]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MTO OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1MTO FirstGlance]. <br>
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</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 3.2&#8491;</td></tr>
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<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=F6P:FRUCTOSE-6-PHOSPHATE'>F6P</scene></td></tr>
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<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1mto FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1mto OCA], [https://pdbe.org/1mto PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1mto RCSB], [https://www.ebi.ac.uk/pdbsum/1mto PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1mto ProSAT]</span></td></tr>
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</table>
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== Function ==
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[https://www.uniprot.org/uniprot/PFKA_GEOSE PFKA_GEOSE] Catalyzes the phosphorylation of D-fructose 6-phosphate to fructose 1,6-bisphosphate by ATP, the first committing step of glycolysis.[HAMAP-Rule:MF_00339]<ref>PMID:8136379</ref>
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== Evolutionary Conservation ==
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[[Image:Consurf_key_small.gif|200px|right]]
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Check<jmol>
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<jmolCheckbox>
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<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/mt/1mto_consurf.spt"</scriptWhenChecked>
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<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked>
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<text>to colour the structure by Evolutionary Conservation</text>
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</jmolCheckbox>
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</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1mto ConSurf].
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<div style="clear:both"></div>
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==About this Structure==
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==See Also==
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1MTO is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Geobacillus_stearothermophilus Geobacillus stearothermophilus] with F6P as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/6-phosphofructokinase 6-phosphofructokinase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.7.1.11 2.7.1.11] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1MTO OCA].
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*[[Phosphofructokinase 3D structures|Phosphofructokinase 3D structures]]
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== References ==
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==Reference==
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<references/>
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Reversible ligand-induced dissociation of a tryptophan-shift mutant of phosphofructokinase from Bacillus stearothermophilus., Riley-Lovingshimer MR, Ronning DR, Sacchettini JC, Reinhart GD, Biochemistry. 2002 Oct 29;41(43):12967-74. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=12390023 12390023]
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__TOC__
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[[Category: 6-phosphofructokinase]]
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</StructureSection>
[[Category: Geobacillus stearothermophilus]]
[[Category: Geobacillus stearothermophilus]]
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[[Category: Single protein]]
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[[Category: Large Structures]]
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[[Category: Reinhart, G.D.]]
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[[Category: Reinhart GD]]
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[[Category: Riley-Lovingshimer, M.R.]]
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[[Category: Riley-Lovingshimer MR]]
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[[Category: Ronning, D.R.]]
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[[Category: Ronning DR]]
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[[Category: Sacchettini, J.C.]]
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[[Category: Sacchettini JC]]
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[[Category: F6P]]
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[[Category: fructose-6-phosphate]]
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[[Category: phosphofructokinase]]
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[[Category: tryptophan-shift]]
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''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 21:41:23 2007''
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Current revision

Crystal structure of a Phosphofructokinase mutant from Bacillus stearothermophilus bound with fructose-6-phosphate

PDB ID 1mto

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