1w3h

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[[Image:1w3h.gif|left|200px]]
[[Image:1w3h.gif|left|200px]]
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{{Structure
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<!--
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|PDB= 1w3h |SIZE=350|CAPTION= <scene name='initialview01'>1w3h</scene>, resolution 1.50&Aring;
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The line below this paragraph, containing "STRUCTURE_1w3h", creates the "Structure Box" on the page.
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|SITE= <scene name='pdbsite=AC1:Egl+Binding+Site+For+Chain+B'>AC1</scene>
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You may change the PDB parameter (which sets the PDB file loaded into the applet)
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|LIGAND= <scene name='pdbligand=CA:CALCIUM+ION'>CA</scene>, <scene name='pdbligand=EDO:1,2-ETHANEDIOL'>EDO</scene>
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or the SCENE parameter (which sets the initial scene displayed when the page is loaded),
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|ACTIVITY= <span class='plainlinks'>[http://en.wikipedia.org/wiki/Endo-1,4-beta-xylanase Endo-1,4-beta-xylanase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.2.1.8 3.2.1.8] </span>
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or leave the SCENE parameter empty for the default display.
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|GENE=
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-->
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|DOMAIN=
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{{STRUCTURE_1w3h| PDB=1w3h | SCENE= }}
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|RELATEDENTRY=
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|RESOURCES=<span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1w3h FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1w3h OCA], [http://www.ebi.ac.uk/pdbsum/1w3h PDBsum], [http://www.rcsb.org/pdb/explore.do?structureId=1w3h RCSB]</span>
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}}
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'''THE 3-DIMENSIONAL STRUCTURE OF A THERMOSTABLE MUTANT OF A XYLANASE (XYN10A) FROM CELLVIBRIO JAPONICUS'''
'''THE 3-DIMENSIONAL STRUCTURE OF A THERMOSTABLE MUTANT OF A XYLANASE (XYN10A) FROM CELLVIBRIO JAPONICUS'''
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[[Category: Taylor, E J.]]
[[Category: Taylor, E J.]]
[[Category: Vincent, F.]]
[[Category: Vincent, F.]]
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[[Category: calcium ion]]
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[[Category: Calcium ion]]
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[[Category: error prone pcr]]
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[[Category: Error prone pcr]]
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[[Category: family 10]]
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[[Category: Family 10]]
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[[Category: glycosyle hydrolase]]
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[[Category: Glycosyle hydrolase]]
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[[Category: hydrolase]]
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[[Category: Hydrolase]]
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[[Category: thermostable]]
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[[Category: Thermostable]]
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[[Category: treble mutant,]]
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[[Category: Treble mutant]]
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[[Category: xylanase]]
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[[Category: Xylanase]]
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sat May 3 13:06:23 2008''
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Mar 31 00:30:40 2008''
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Revision as of 10:06, 3 May 2008

Image:1w3h.gif

Template:STRUCTURE 1w3h

THE 3-DIMENSIONAL STRUCTURE OF A THERMOSTABLE MUTANT OF A XYLANASE (XYN10A) FROM CELLVIBRIO JAPONICUS


Overview

Metal ions such as calcium often play a key role in protein thermostability. The inclusion of metal ions in industrial processes is, however, problematic. Thus, the evolution of enzymes that display enhanced stability, which is not reliant on divalent metals, is an important biotechnological goal. Here we have used forced protein evolution to interrogate whether the stabilizing effect of calcium in an industrially relevant enzyme can be replaced with amino acid substitutions. Our study has focused on the GH10 xylanase CjXyn10A from Cellvibrio japonicus, which contains an extended calcium binding loop that confers proteinase resistance and thermostability. Three rounds of error-prone PCR and selection identified a treble mutant, D262N/A80T/R347C, which in the absence of calcium is more thermostable than wild type CjXyn10A bound to the divalent metal. D262N influences the properties of the calcium binding site, A80T fills a cavity in the enzyme, increasing the number of hydrogen bonds and van der Waals interactions, and the R347C mutation introduces a disulfide bond that decreases the free energy of the unfolded enzyme. A derivative of CjXyn10A (CfCjXyn10A) in which the calcium binding loop has been replaced with a much shorter loop from Cellulomonas fimi CfXyn10A was also subjected to forced protein evolution to select for thermostablizing mutations. Two amino acid substitutions within the introduced loop and the A80T mutation increased the thermostability of the enzyme. This study demonstrates how forced protein evolution can be used to introduce enhanced stability into industrially relevant enzymes while removing calcium as a major stability determinant.

About this Structure

1W3H is a Single protein structure of sequence from Cellvibrio japonicus. Full crystallographic information is available from OCA.

Reference

The use of forced protein evolution to investigate and improve stability of family 10 xylanases. The production of Ca2+-independent stable xylanases., Andrews SR, Taylor EJ, Pell G, Vincent F, Ducros VM, Davies GJ, Lakey JH, Gilbert HJ, J Biol Chem. 2004 Dec 24;279(52):54369-79. Epub 2004 Sep 27. PMID:15452124 Page seeded by OCA on Sat May 3 13:06:23 2008

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