4pc8: Difference between revisions
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The | ==Structure-based protein engineering efforts on the scaffold of a monomeric triosephosphate isomerase yielding a sugar isomerase== | ||
<StructureSection load='4pc8' size='340' side='right'caption='[[4pc8]], [[Resolution|resolution]] 1.55Å' scene=''> | |||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[4pc8]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Trypanosoma_brucei_brucei Trypanosoma brucei brucei]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4PC8 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4PC8 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]] 1.55Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=GOA:GLYCOLIC+ACID'>GOA</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=4pc8 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4pc8 OCA], [https://pdbe.org/4pc8 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4pc8 RCSB], [https://www.ebi.ac.uk/pdbsum/4pc8 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4pc8 ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/TPIS_TRYBB TPIS_TRYBB] | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
The crystal structures are described of two variants of A-TIM: Ma18 (2.7 A resolution) and Ma21 (1.55 A resolution). A-TIM is a monomeric loop-deletion variant of triosephosphate isomerase (TIM) which has lost the TIM catalytic properties. Ma18 and Ma21 were identified after extensive directed-evolution selection experiments using an Escherichia coli L-arabinose isomerase knockout strain expressing a randomly mutated A-TIM gene. These variants facilitate better growth of the Escherichia coli selection strain in medium supplemented with 40 mM L-arabinose. Ma18 and Ma21 differ from A-TIM by four and one point mutations, respectively. Ma18 and Ma21 are more stable proteins than A-TIM, as judged from CD melting experiments. Like A-TIM, both proteins are monomeric in solution. In the Ma18 crystal structure loop 6 is open and in the Ma21 crystal structure loop 6 is closed, being stabilized by a bound glycolate molecule. The crystal structures show only small differences in the active site compared with A-TIM. In the case of Ma21 it is observed that the point mutation (Q65L) contributes to small structural rearrangements near Asn11 of loop 1, which correlate with different ligand-binding properties such as a loss of citrate binding in the active site. The Ma21 structure also shows that its Leu65 side chain is involved in van der Waals interactions with neighbouring hydrophobic side-chain moieties, correlating with its increased stability. The experimental data suggest that the increased stability and solubility properties of Ma21 and Ma18 compared with A-TIM cause better growth of the selection strain when coexpressing Ma21 and Ma18 instead of A-TIM. | |||
Crystal structures of two monomeric triosephosphate isomerase variants identified via a directed-evolution protocol selecting for L-arabinose isomerase activity.,Krause M, Kiema TR, Neubauer P, Wierenga RK Acta Crystallogr F Struct Biol Commun. 2016 Jun 1;72(Pt 6):490-9. doi:, 10.1107/S2053230X16007548. Epub 2016 May 23. PMID:27303904<ref>PMID:27303904</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
<div class="pdbe-citations 4pc8" style="background-color:#fffaf0;"></div> | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
[[Category: Large Structures]] | |||
[[Category: Trypanosoma brucei brucei]] | |||
[[Category: Krause M]] | |||
[[Category: Neubauer P]] | |||
[[Category: Wierenga RK]] | |||
Latest revision as of 12:20, 15 November 2023
Structure-based protein engineering efforts on the scaffold of a monomeric triosephosphate isomerase yielding a sugar isomeraseStructure-based protein engineering efforts on the scaffold of a monomeric triosephosphate isomerase yielding a sugar isomerase
Structural highlights
FunctionPublication Abstract from PubMedThe crystal structures are described of two variants of A-TIM: Ma18 (2.7 A resolution) and Ma21 (1.55 A resolution). A-TIM is a monomeric loop-deletion variant of triosephosphate isomerase (TIM) which has lost the TIM catalytic properties. Ma18 and Ma21 were identified after extensive directed-evolution selection experiments using an Escherichia coli L-arabinose isomerase knockout strain expressing a randomly mutated A-TIM gene. These variants facilitate better growth of the Escherichia coli selection strain in medium supplemented with 40 mM L-arabinose. Ma18 and Ma21 differ from A-TIM by four and one point mutations, respectively. Ma18 and Ma21 are more stable proteins than A-TIM, as judged from CD melting experiments. Like A-TIM, both proteins are monomeric in solution. In the Ma18 crystal structure loop 6 is open and in the Ma21 crystal structure loop 6 is closed, being stabilized by a bound glycolate molecule. The crystal structures show only small differences in the active site compared with A-TIM. In the case of Ma21 it is observed that the point mutation (Q65L) contributes to small structural rearrangements near Asn11 of loop 1, which correlate with different ligand-binding properties such as a loss of citrate binding in the active site. The Ma21 structure also shows that its Leu65 side chain is involved in van der Waals interactions with neighbouring hydrophobic side-chain moieties, correlating with its increased stability. The experimental data suggest that the increased stability and solubility properties of Ma21 and Ma18 compared with A-TIM cause better growth of the selection strain when coexpressing Ma21 and Ma18 instead of A-TIM. Crystal structures of two monomeric triosephosphate isomerase variants identified via a directed-evolution protocol selecting for L-arabinose isomerase activity.,Krause M, Kiema TR, Neubauer P, Wierenga RK Acta Crystallogr F Struct Biol Commun. 2016 Jun 1;72(Pt 6):490-9. doi:, 10.1107/S2053230X16007548. Epub 2016 May 23. PMID:27303904[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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