1knd
From Proteopedia
(New page: 200px<br /><applet load="1knd" size="450" color="white" frame="true" align="right" spinBox="true" caption="1knd, resolution 1.9Å" /> '''Crystal Structure of ...) |
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| - | [[Image:1knd.gif|left|200px]]<br /><applet load="1knd" size=" | + | [[Image:1knd.gif|left|200px]]<br /><applet load="1knd" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1knd, resolution 1.9Å" /> | caption="1knd, resolution 1.9Å" /> | ||
'''Crystal Structure of 2,3-dihydroxybiphenyl 1,2-dioxygenase Complexed with Catechol under Anaerobic Condition'''<br /> | '''Crystal Structure of 2,3-dihydroxybiphenyl 1,2-dioxygenase Complexed with Catechol under Anaerobic Condition'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The steady-state cleavage of catechols by 2,3-dihydroxybiphenyl 1, 2-dioxygenase (DHBD), the extradiol dioxygenase of the biphenyl | + | The steady-state cleavage of catechols by 2,3-dihydroxybiphenyl 1, 2-dioxygenase (DHBD), the extradiol dioxygenase of the biphenyl biodegradation pathway, was investigated using a highly active, anaerobically purified preparation of enzyme. The kinetic data obtained using 2,3-dihydroxybiphenyl (DHB) fit a compulsory order ternary complex mechanism in which substrate inhibition occurs. The Km for dioxygen was 1280 +/- 70 microM, which is at least 2 orders of magnitude higher than that reported for catechol 2,3-dioxygenases. Km and Kd for DHB were 22 +/- 2 and 8 +/- 1 microM, respectively. DHBD was subject to reversible substrate inhibition and mechanism-based inactivation. In air-saturated buffer, the partition ratios of catecholic substrates substituted at C-3 were inversely related to their apparent specificity constants. Small organic molecules that stabilized DHBD most effectively also inhibited the cleavage reaction most strongly. The steady-state kinetic data and crystallographic results suggest that the stabilization and inhibition are due to specific interactions between the organic molecule and the active site of the enzyme. t-Butanol stabilized the enzyme and inhibited the cleavage of DHB in a mixed fashion, consistent with the distinct binding sites occupied by t-butanol in the crystal structures of the substrate-free form of the enzyme and the enzyme-DHB complex. In contrast, crystal structures of complexes with catechol and 3-methylcatechol revealed relationships between the binding of these smaller substrates and t-butanol that are consistent with the observed competitive inhibition. |
==About this Structure== | ==About this Structure== | ||
| - | 1KND is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Burkholderia_cepacia Burkholderia cepacia] with FE2, CAQ and TBU as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Biphenyl-2,3-diol_1,2-dioxygenase Biphenyl-2,3-diol 1,2-dioxygenase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.13.11.39 1.13.11.39] Full crystallographic information is available from [http:// | + | 1KND is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Burkholderia_cepacia Burkholderia cepacia] with <scene name='pdbligand=FE2:'>FE2</scene>, <scene name='pdbligand=CAQ:'>CAQ</scene> and <scene name='pdbligand=TBU:'>TBU</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Biphenyl-2,3-diol_1,2-dioxygenase Biphenyl-2,3-diol 1,2-dioxygenase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.13.11.39 1.13.11.39] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1KND OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Burkholderia cepacia]] | [[Category: Burkholderia cepacia]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Bolin, J | + | [[Category: Bolin, J T.]] |
[[Category: Han, S.]] | [[Category: Han, S.]] | ||
[[Category: CAQ]] | [[Category: CAQ]] | ||
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[[Category: oxidoreductase]] | [[Category: oxidoreductase]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 13:35:56 2008'' |
Revision as of 11:36, 21 February 2008
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Crystal Structure of 2,3-dihydroxybiphenyl 1,2-dioxygenase Complexed with Catechol under Anaerobic Condition
Overview
The steady-state cleavage of catechols by 2,3-dihydroxybiphenyl 1, 2-dioxygenase (DHBD), the extradiol dioxygenase of the biphenyl biodegradation pathway, was investigated using a highly active, anaerobically purified preparation of enzyme. The kinetic data obtained using 2,3-dihydroxybiphenyl (DHB) fit a compulsory order ternary complex mechanism in which substrate inhibition occurs. The Km for dioxygen was 1280 +/- 70 microM, which is at least 2 orders of magnitude higher than that reported for catechol 2,3-dioxygenases. Km and Kd for DHB were 22 +/- 2 and 8 +/- 1 microM, respectively. DHBD was subject to reversible substrate inhibition and mechanism-based inactivation. In air-saturated buffer, the partition ratios of catecholic substrates substituted at C-3 were inversely related to their apparent specificity constants. Small organic molecules that stabilized DHBD most effectively also inhibited the cleavage reaction most strongly. The steady-state kinetic data and crystallographic results suggest that the stabilization and inhibition are due to specific interactions between the organic molecule and the active site of the enzyme. t-Butanol stabilized the enzyme and inhibited the cleavage of DHB in a mixed fashion, consistent with the distinct binding sites occupied by t-butanol in the crystal structures of the substrate-free form of the enzyme and the enzyme-DHB complex. In contrast, crystal structures of complexes with catechol and 3-methylcatechol revealed relationships between the binding of these smaller substrates and t-butanol that are consistent with the observed competitive inhibition.
About this Structure
1KND is a Single protein structure of sequence from Burkholderia cepacia with , and as ligands. Active as Biphenyl-2,3-diol 1,2-dioxygenase, with EC number 1.13.11.39 Full crystallographic information is available from OCA.
Reference
Molecular basis for the stabilization and inhibition of 2, 3-dihydroxybiphenyl 1,2-dioxygenase by t-butanol., Vaillancourt FH, Han S, Fortin PD, Bolin JT, Eltis LD, J Biol Chem. 1998 Dec 25;273(52):34887-95. PMID:9857017
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