1yoo
From Proteopedia
(New page: 200px<br /><applet load="1yoo" size="450" color="white" frame="true" align="right" spinBox="true" caption="1yoo, resolution 2.40Å" /> '''ASPARTATE AMINOTRANS...) |
|||
| Line 1: | Line 1: | ||
| - | [[Image:1yoo.jpg|left|200px]]<br /><applet load="1yoo" size=" | + | [[Image:1yoo.jpg|left|200px]]<br /><applet load="1yoo" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1yoo, resolution 2.40Å" /> | caption="1yoo, resolution 2.40Å" /> | ||
'''ASPARTATE AMINOTRANSFERASE MUTANT ATB17 WITH ISOVALERIC ACID'''<br /> | '''ASPARTATE AMINOTRANSFERASE MUTANT ATB17 WITH ISOVALERIC ACID'''<br /> | ||
==Overview== | ==Overview== | ||
| - | Directed evolution was used to change the substrate specificity of | + | Directed evolution was used to change the substrate specificity of aspartate aminotransferase. A mutant enzyme with 17 amino acid substitutions was generated that shows a 2.1 x 10(6)-fold increase in the catalytic efficiency (kcat/Km) for a non-native substrate, valine. The absorption spectrum of the bound coenzyme, pyridoxal 5'-phosphate, is also changed significantly by the mutations. Interestingly, only one of the 17 residues appears to be able to contact the substrate, and none of them interact with the coenzyme. The three-dimensional structure of the mutant enzyme complexed with a valine analog, isovalerate (determined to 2.4-A resolution by x-ray crystallography), provides insights into how the mutations affect substrate binding. The active site is remodeled; the subunit interface is altered, and the enzyme domain that encloses the substrate is shifted by the mutations. The present results demonstrate clearly the importance of the cumulative effects of residues remote from the active site and represent a new line of approach to the redesign of enzyme activity. |
==About this Structure== | ==About this Structure== | ||
| - | 1YOO is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with PLP and IVA as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Aspartate_transaminase Aspartate transaminase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.6.1.1 2.6.1.1] Full crystallographic information is available from [http:// | + | 1YOO is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with <scene name='pdbligand=PLP:'>PLP</scene> and <scene name='pdbligand=IVA:'>IVA</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Aspartate_transaminase Aspartate transaminase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.6.1.1 2.6.1.1] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1YOO OCA]. |
==Reference== | ==Reference== | ||
| Line 22: | Line 22: | ||
[[Category: aminotransferase]] | [[Category: aminotransferase]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:07:37 2008'' |
Revision as of 14:07, 21 February 2008
|
ASPARTATE AMINOTRANSFERASE MUTANT ATB17 WITH ISOVALERIC ACID
Overview
Directed evolution was used to change the substrate specificity of aspartate aminotransferase. A mutant enzyme with 17 amino acid substitutions was generated that shows a 2.1 x 10(6)-fold increase in the catalytic efficiency (kcat/Km) for a non-native substrate, valine. The absorption spectrum of the bound coenzyme, pyridoxal 5'-phosphate, is also changed significantly by the mutations. Interestingly, only one of the 17 residues appears to be able to contact the substrate, and none of them interact with the coenzyme. The three-dimensional structure of the mutant enzyme complexed with a valine analog, isovalerate (determined to 2.4-A resolution by x-ray crystallography), provides insights into how the mutations affect substrate binding. The active site is remodeled; the subunit interface is altered, and the enzyme domain that encloses the substrate is shifted by the mutations. The present results demonstrate clearly the importance of the cumulative effects of residues remote from the active site and represent a new line of approach to the redesign of enzyme activity.
About this Structure
1YOO is a Single protein structure of sequence from Escherichia coli with and as ligands. Active as Aspartate transaminase, with EC number 2.6.1.1 Full crystallographic information is available from OCA.
Reference
Redesigning the substrate specificity of an enzyme by cumulative effects of the mutations of non-active site residues., Oue S, Okamoto A, Yano T, Kagamiyama H, J Biol Chem. 1999 Jan 22;274(4):2344-9. PMID:9891001
Page seeded by OCA on Thu Feb 21 16:07:37 2008
