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New page: left|200px<br /><applet load="1t4m" size="450" color="white" frame="true" align="right" spinBox="true" caption="1t4m, resolution 2.00Å" /> '''STRUCTURE OF A THERM...
 
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[[Image:1t4m.jpg|left|200px]]<br /><applet load="1t4m" size="450" color="white" frame="true" align="right" spinBox="true"
caption="1t4m, resolution 2.00&Aring;" />
'''STRUCTURE OF A THERMOSTABLE DOUBLE MUTANT OF BACILLUS SUBTILIS LIPASE OBTAINED THROUGH DIRECTED EVOLUTION'''<br />


==Overview==
==STRUCTURE OF A THERMOSTABLE DOUBLE MUTANT OF BACILLUS SUBTILIS LIPASE OBTAINED THROUGH DIRECTED EVOLUTION==
Variation in gene sequences generated by directed evolution approaches, often does not assure a minimalist design for obtaining a desired property, in proteins. While screening for enhanced thermostability, structural, information was utilized in selecting mutations that are generated by, error-prone PCR. By this approach we have increased the half-life of, denaturation by 300-fold compared to the wild-type Bacillus subtilis, lipase through three point mutations generated by only two cycles of, error-prone PCR. At lower temperatures the activity parameters of the, thermostable mutants are unaltered. High-resolution crystal structures of, the mutants show subtle changes, which include stacking of tyrosine, residues, peptide plane flipping and a better anchoring of the terminus, that challenge rational design and explain the structural basis for, enhanced thermostability. The approach may offer an efficient and, minimalist solution for the enhancement of a desired property of a, protein.
<StructureSection load='1t4m' size='340' side='right'caption='[[1t4m]], [[Resolution|resolution]] 2.00&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[1t4m]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Bacillus_subtilis Bacillus subtilis]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1T4M OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1T4M 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&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=K:POTASSIUM+ION'>K</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=1t4m FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1t4m OCA], [https://pdbe.org/1t4m PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1t4m RCSB], [https://www.ebi.ac.uk/pdbsum/1t4m PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1t4m ProSAT]</span></td></tr>
</table>
== Function ==
[https://www.uniprot.org/uniprot/ESTA_BACSU ESTA_BACSU] Active toward p-nitrophenyl esters and triacylglycerides with a marked preference for esters with C8 acyl groups.<ref>PMID:8396026</ref>
== 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/t4/1t4m_consurf.spt"</scriptWhenChecked>
    <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.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=1t4m ConSurf].
<div style="clear:both"></div>
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
Variation in gene sequences generated by directed evolution approaches often does not assure a minimalist design for obtaining a desired property in proteins. While screening for enhanced thermostability, structural information was utilized in selecting mutations that are generated by error-prone PCR. By this approach we have increased the half-life of denaturation by 300-fold compared to the wild-type Bacillus subtilis lipase through three point mutations generated by only two cycles of error-prone PCR. At lower temperatures the activity parameters of the thermostable mutants are unaltered. High-resolution crystal structures of the mutants show subtle changes, which include stacking of tyrosine residues, peptide plane flipping and a better anchoring of the terminus, that challenge rational design and explain the structural basis for enhanced thermostability. The approach may offer an efficient and minimalist solution for the enhancement of a desired property of a protein.


==About this Structure==
Structural basis of selection and thermostability of laboratory evolved Bacillus subtilis lipase.,Acharya P, Rajakumara E, Sankaranarayanan R, Rao NM J Mol Biol. 2004 Aug 27;341(5):1271-81. PMID:15321721<ref>PMID:15321721</ref>
1T4M is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Bacillus_subtilis Bacillus subtilis] with K as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Triacylglycerol_lipase Triacylglycerol lipase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.1.1.3 3.1.1.3] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1T4M OCA].


==Reference==
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
Structural basis of selection and thermostability of laboratory evolved Bacillus subtilis lipase., Acharya P, Rajakumara E, Sankaranarayanan R, Rao NM, J Mol Biol. 2004 Aug 27;341(5):1271-81. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=15321721 15321721]
</div>
<div class="pdbe-citations 1t4m" style="background-color:#fffaf0;"></div>
 
==See Also==
*[[Lipase 3D Structures|Lipase 3D Structures]]
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Bacillus subtilis]]
[[Category: Bacillus subtilis]]
[[Category: Single protein]]
[[Category: Large Structures]]
[[Category: Triacylglycerol lipase]]
[[Category: Rajakumara E]]
[[Category: Rajakumara, E.]]
[[Category: Sankaranarayanan R]]
[[Category: Sankaranarayanan, R.]]
[[Category: K]]
[[Category: alpha/beta hydrolase]]
 
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