1oa2: Difference between revisions
New page: left|200px<br /> <applet load="1oa2" size="450" color="white" frame="true" align="right" spinBox="true" caption="1oa2, resolution 1.5Å" /> '''COMPARISON OF FAMILY... |
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== | ==Comparison of Family 12 Glycoside Hydrolases and Recruited Substitutions Important for Thermal Stability== | ||
As part of a program to discover improved glycoside hydrolase family 12 | <StructureSection load='1oa2' size='340' side='right'caption='[[1oa2]], [[Resolution|resolution]] 1.50Å' scene=''> | ||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[1oa2]] is a 6 chain structure with sequence from [https://en.wikipedia.org/wiki/Trichoderma_reesei Trichoderma reesei]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1OA2 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1OA2 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]] 1.5Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><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=1oa2 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1oa2 OCA], [https://pdbe.org/1oa2 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1oa2 RCSB], [https://www.ebi.ac.uk/pdbsum/1oa2 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1oa2 ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/O00095_HYPJE O00095_HYPJE] | |||
== 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/oa/1oa2_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=1oa2 ConSurf]. | |||
<div style="clear:both"></div> | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
As part of a program to discover improved glycoside hydrolase family 12 (GH 12) endoglucanases, we have studied the biochemical diversity of several GH 12 homologs. The H. schweinitzii Cel12A enzyme differs from the T. reesei Cel12A enzyme by only 14 amino acids (93% sequence identity), but is much less thermally stable. The bacterial Cel12A enzyme from S. sp. 11AG8 shares only 28% sequence identity to the T. reesei enzyme, and is much more thermally stable. Each of the 14 sequence differences from H. schweinitzii Cel12A were introduced in T. reesei Cel12A to determine the effect of these amino acid substitutions on enzyme stability. Several of the T. reesei Cel12A variants were found to have increased stability, and the differences in apparent midpoint of thermal denaturation (T(m)) ranged from a 2.5 degrees C increase to a 4.0 degrees C decrease. The least stable recruitment from H. schweinitzii Cel12A was A35S. Consequently, the A35V substitution was recruited from the more stable S. sp. 11AG8 Cel12A and this T. reesei Cel12A variant was found to have a T(m) 7.7 degrees C higher than wild type. Thus, the buried residue at position 35 was shown to be of critical importance for thermal stability in this structural family. There was a ninefold range in the specific activities of the Cel12 homologs on o-NPC. The most and least stable T. reesei Cel12A variants, A35V and A35S, respectively, were fully active. Because of their thermal tolerance, S. sp. 11AG8 Cel12A and T. reesei Cel12A variant A35V showed a continual increase in activity over the temperature range of 25 degrees C to 60 degrees C, whereas the less stable enzymes T. reesei Cel12A wild type and the destabilized A35S variant, and H. schweinitzii Cel12A showed a decrease in activity at the highest temperatures. The crystal structures of the H. schweinitzii, S. sp. 11AG8, and T. reesei A35V Cel12A enzymes have been determined and compared with the wild-type T. reesei Cel12A enzyme. All of the structures have similar Calpha traces, but provide detailed insight into the nature of the stability differences. These results are an example of the power of homolog recruitment as a method for identifying residues important for stability. | |||
Comparison of family 12 glycoside hydrolases and recruited substitutions important for thermal stability.,Sandgren M, Gualfetti PJ, Shaw A, Gross LS, Saldajeno M, Day AG, Jones TA, Mitchinson C Protein Sci. 2003 Apr;12(4):848-60. PMID:12649442<ref>PMID:12649442</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
[[Category: | <div class="pdbe-citations 1oa2" style="background-color:#fffaf0;"></div> | ||
==See Also== | |||
*[[Glucanase 3D structures|Glucanase 3D structures]] | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
[[Category: Large Structures]] | |||
[[Category: Trichoderma reesei]] | [[Category: Trichoderma reesei]] | ||
[[Category: Day | [[Category: Day AG]] | ||
[[Category: Gross | [[Category: Gross LS]] | ||
[[Category: Gualfetti | [[Category: Gualfetti PJ]] | ||
[[Category: Jones | [[Category: Jones TA]] | ||
[[Category: Mitchinson | [[Category: Mitchinson C]] | ||
[[Category: Saldajeno | [[Category: Saldajeno M]] | ||
[[Category: Sandgren | [[Category: Sandgren M]] | ||
[[Category: Shaw | [[Category: Shaw A]] | ||
Latest revision as of 11:41, 6 November 2024
Comparison of Family 12 Glycoside Hydrolases and Recruited Substitutions Important for Thermal StabilityComparison of Family 12 Glycoside Hydrolases and Recruited Substitutions Important for Thermal Stability
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 PubMedAs part of a program to discover improved glycoside hydrolase family 12 (GH 12) endoglucanases, we have studied the biochemical diversity of several GH 12 homologs. The H. schweinitzii Cel12A enzyme differs from the T. reesei Cel12A enzyme by only 14 amino acids (93% sequence identity), but is much less thermally stable. The bacterial Cel12A enzyme from S. sp. 11AG8 shares only 28% sequence identity to the T. reesei enzyme, and is much more thermally stable. Each of the 14 sequence differences from H. schweinitzii Cel12A were introduced in T. reesei Cel12A to determine the effect of these amino acid substitutions on enzyme stability. Several of the T. reesei Cel12A variants were found to have increased stability, and the differences in apparent midpoint of thermal denaturation (T(m)) ranged from a 2.5 degrees C increase to a 4.0 degrees C decrease. The least stable recruitment from H. schweinitzii Cel12A was A35S. Consequently, the A35V substitution was recruited from the more stable S. sp. 11AG8 Cel12A and this T. reesei Cel12A variant was found to have a T(m) 7.7 degrees C higher than wild type. Thus, the buried residue at position 35 was shown to be of critical importance for thermal stability in this structural family. There was a ninefold range in the specific activities of the Cel12 homologs on o-NPC. The most and least stable T. reesei Cel12A variants, A35V and A35S, respectively, were fully active. Because of their thermal tolerance, S. sp. 11AG8 Cel12A and T. reesei Cel12A variant A35V showed a continual increase in activity over the temperature range of 25 degrees C to 60 degrees C, whereas the less stable enzymes T. reesei Cel12A wild type and the destabilized A35S variant, and H. schweinitzii Cel12A showed a decrease in activity at the highest temperatures. The crystal structures of the H. schweinitzii, S. sp. 11AG8, and T. reesei A35V Cel12A enzymes have been determined and compared with the wild-type T. reesei Cel12A enzyme. All of the structures have similar Calpha traces, but provide detailed insight into the nature of the stability differences. These results are an example of the power of homolog recruitment as a method for identifying residues important for stability. Comparison of family 12 glycoside hydrolases and recruited substitutions important for thermal stability.,Sandgren M, Gualfetti PJ, Shaw A, Gross LS, Saldajeno M, Day AG, Jones TA, Mitchinson C Protein Sci. 2003 Apr;12(4):848-60. PMID:12649442[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences |
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