1kb3: Difference between revisions
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< | ==Three Dimensional Structure Analysis of the R195A Variant of Human Pancreatic Alpha Amylase== | ||
<StructureSection load='1kb3' size='340' side='right'caption='[[1kb3]], [[Resolution|resolution]] 2.10Å' scene=''> | |||
You may | == Structural highlights == | ||
<table><tr><td colspan='2'>[[1kb3]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1KB3 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1KB3 FirstGlance]. <br> | |||
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.1Å</td></tr> | |||
-- | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CA:CALCIUM+ION'>CA</scene>, <scene name='pdbligand=NAG:N-ACETYL-D-GLUCOSAMINE'>NAG</scene>, <scene name='pdbligand=PCA:PYROGLUTAMIC+ACID'>PCA</scene></td></tr> | ||
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1kb3 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1kb3 OCA], [https://pdbe.org/1kb3 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1kb3 RCSB], [https://www.ebi.ac.uk/pdbsum/1kb3 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1kb3 ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/AMYP_HUMAN AMYP_HUMAN] | |||
== Evolutionary Conservation == | |||
[[Image:Consurf_key_small.gif|200px|right]] | |||
Check<jmol> | |||
<jmolCheckbox> | |||
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/kb/1kb3_consurf.spt"</scriptWhenChecked> | |||
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview03.spt</scriptWhenUnchecked> | |||
<text>to colour the structure by Evolutionary Conservation</text> | |||
</jmolCheckbox> | |||
</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=1kb3 ConSurf]. | |||
<div style="clear:both"></div> | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Human pancreatic alpha-amylase (HPA) is a member of the alpha-amylase family involved in the degradation of starch. Some members of this family, including HPA, require chloride for maximal activity. To determine the mechanism of chloride activation, a series of mutants (R195A, R195Q, N298S, R337A, and R337Q) were made in which residues in the chloride ion binding site were replaced. Mutations in this binding site were found to severely affect the ability of HPA to bind chloride ions with no binding detected for the R195 and R337 mutant enzymes. X-ray crystallographic analysis revealed that these mutations did not result in significant structural changes. However, the introduction of these mutations did alter the kinetic properties of the enzyme. Mutations to residue R195 resulted in a 20-450-fold decrease in the activity of the enzyme toward starch and shifted the pH optimum to a more basic pH. Interestingly, replacement of R337 with a nonbasic amino acid resulted in an alpha-amylase that no longer required chloride for catalysis and has a pH profile similar to that of wild-type HPA. In contrast, a mutation at residue N298 resulted in an enzyme that had much lower binding affinity for chloride but still required chloride for maximal activity. We propose that the chloride is required to increase the pK(a) of the acid/base catalyst, E233, which would otherwise be lower due to the presence of R337, a positively charged residue. | |||
Probing the role of the chloride ion in the mechanism of human pancreatic alpha-amylase.,Numao S, Maurus R, Sidhu G, Wang Y, Overall CM, Brayer GD, Withers SG Biochemistry. 2002 Jan 8;41(1):215-25. PMID:11772019<ref>PMID:11772019</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
<div class="pdbe-citations 1kb3" style="background-color:#fffaf0;"></div> | |||
==See Also== | |||
*[[Amylase 3D structures|Amylase 3D structures]] | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
== | |||
== | |||
[[Category: Homo sapiens]] | [[Category: Homo sapiens]] | ||
[[Category: | [[Category: Large Structures]] | ||
[[Category: Brayer | [[Category: Brayer GD]] | ||
[[Category: Maurus | [[Category: Maurus R]] | ||
[[Category: Numao | [[Category: Numao S]] | ||
[[Category: Overall | [[Category: Overall CM]] | ||
[[Category: Sidhu | [[Category: Sidhu G]] | ||
[[Category: Wang | [[Category: Wang Y]] | ||
[[Category: Withers | [[Category: Withers SG]] | ||
Latest revision as of 11:34, 6 November 2024
Three Dimensional Structure Analysis of the R195A Variant of Human Pancreatic Alpha AmylaseThree Dimensional Structure Analysis of the R195A Variant of Human Pancreatic Alpha Amylase
Structural highlights
FunctionEvolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedHuman pancreatic alpha-amylase (HPA) is a member of the alpha-amylase family involved in the degradation of starch. Some members of this family, including HPA, require chloride for maximal activity. To determine the mechanism of chloride activation, a series of mutants (R195A, R195Q, N298S, R337A, and R337Q) were made in which residues in the chloride ion binding site were replaced. Mutations in this binding site were found to severely affect the ability of HPA to bind chloride ions with no binding detected for the R195 and R337 mutant enzymes. X-ray crystallographic analysis revealed that these mutations did not result in significant structural changes. However, the introduction of these mutations did alter the kinetic properties of the enzyme. Mutations to residue R195 resulted in a 20-450-fold decrease in the activity of the enzyme toward starch and shifted the pH optimum to a more basic pH. Interestingly, replacement of R337 with a nonbasic amino acid resulted in an alpha-amylase that no longer required chloride for catalysis and has a pH profile similar to that of wild-type HPA. In contrast, a mutation at residue N298 resulted in an enzyme that had much lower binding affinity for chloride but still required chloride for maximal activity. We propose that the chloride is required to increase the pK(a) of the acid/base catalyst, E233, which would otherwise be lower due to the presence of R337, a positively charged residue. Probing the role of the chloride ion in the mechanism of human pancreatic alpha-amylase.,Numao S, Maurus R, Sidhu G, Wang Y, Overall CM, Brayer GD, Withers SG Biochemistry. 2002 Jan 8;41(1):215-25. PMID:11772019[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences |
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