1uio
From Proteopedia
(New page: 200px<br /><applet load="1uio" size="450" color="white" frame="true" align="right" spinBox="true" caption="1uio, resolution 2.4Å" /> '''ADENOSINE DEAMINASE (...) |
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| - | [[Image:1uio.jpg|left|200px]]<br /><applet load="1uio" size=" | + | [[Image:1uio.jpg|left|200px]]<br /><applet load="1uio" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1uio, resolution 2.4Å" /> | caption="1uio, resolution 2.4Å" /> | ||
'''ADENOSINE DEAMINASE (HIS 238 ALA MUTANT)'''<br /> | '''ADENOSINE DEAMINASE (HIS 238 ALA MUTANT)'''<br /> | ||
==Overview== | ==Overview== | ||
| - | His 238, a conserved amino acid located in hydrogen-bonding distance from | + | His 238, a conserved amino acid located in hydrogen-bonding distance from C-6 of the substrate in the active site of murine adenosine deaminase (mADA) and postulated to play an important role in catalysis, was altered into an alanine, a glutamate, and an arginine using site-directed mutagenesis. The Ala and Glu substitutions did not result in changes of the secondary or tertiary structure, while the Arg mutation caused local perturbations in tertiary structure and quenched the emission of one or more enzyme tryptophans. Neither the Glu or Arg mutations affected substrate binding affinity. By contrast, the Ala mutation enhanced substrate and inhibitor binding by 20-fold. The most inactive of the mutants, Glu 238, had a kcat/K(m) 4 x 10(-6) lower than the wild-type value, suggesting that a positive charge on His 238 is important for proper catalytic function. The Ala 238 mutant was the most active ADA, with a kcat/K(m) 2 x 10(-3) lower than the wild-type value. NMR spectroscopy and crystallography revealed that this mutant is able to catalyze hydration of purine riboside, a ground-state analog of the reaction. These results collectively show that His 238 is not required for formation of the hydroxylate used in the deamination and may instead have an important electrostatic role. |
==About this Structure== | ==About this Structure== | ||
| - | 1UIO is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Mus_musculus Mus musculus] with ZN and HPR as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Adenosine_deaminase Adenosine deaminase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.4.4 3.5.4.4] Full crystallographic information is available from [http:// | + | 1UIO is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Mus_musculus Mus musculus] with <scene name='pdbligand=ZN:'>ZN</scene> and <scene name='pdbligand=HPR:'>HPR</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Adenosine_deaminase Adenosine deaminase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.4.4 3.5.4.4] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1UIO OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Mus musculus]] | [[Category: Mus musculus]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Quiocho, F | + | [[Category: Quiocho, F A.]] |
| - | [[Category: Wilson, D | + | [[Category: Wilson, D K.]] |
[[Category: HPR]] | [[Category: HPR]] | ||
[[Category: ZN]] | [[Category: ZN]] | ||
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[[Category: zinc cofactor tim barrel]] | [[Category: zinc cofactor tim barrel]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 15:25:02 2008'' |
Revision as of 13:25, 21 February 2008
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ADENOSINE DEAMINASE (HIS 238 ALA MUTANT)
Overview
His 238, a conserved amino acid located in hydrogen-bonding distance from C-6 of the substrate in the active site of murine adenosine deaminase (mADA) and postulated to play an important role in catalysis, was altered into an alanine, a glutamate, and an arginine using site-directed mutagenesis. The Ala and Glu substitutions did not result in changes of the secondary or tertiary structure, while the Arg mutation caused local perturbations in tertiary structure and quenched the emission of one or more enzyme tryptophans. Neither the Glu or Arg mutations affected substrate binding affinity. By contrast, the Ala mutation enhanced substrate and inhibitor binding by 20-fold. The most inactive of the mutants, Glu 238, had a kcat/K(m) 4 x 10(-6) lower than the wild-type value, suggesting that a positive charge on His 238 is important for proper catalytic function. The Ala 238 mutant was the most active ADA, with a kcat/K(m) 2 x 10(-3) lower than the wild-type value. NMR spectroscopy and crystallography revealed that this mutant is able to catalyze hydration of purine riboside, a ground-state analog of the reaction. These results collectively show that His 238 is not required for formation of the hydroxylate used in the deamination and may instead have an important electrostatic role.
About this Structure
1UIO is a Single protein structure of sequence from Mus musculus with and as ligands. Active as Adenosine deaminase, with EC number 3.5.4.4 Full crystallographic information is available from OCA.
Reference
Site-directed mutagenesis of histidine 238 in mouse adenosine deaminase: substitution of histidine 238 does not impede hydroxylate formation., Sideraki V, Wilson DK, Kurz LC, Quiocho FA, Rudolph FB, Biochemistry. 1996 Nov 26;35(47):15019-28. PMID:8942668
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