2ddf
From Proteopedia
(New page: 200px<br /> <applet load="2ddf" size="450" color="white" frame="true" align="right" spinBox="true" caption="2ddf, resolution 1.700Å" /> '''Crystal structure ...) |
|||
| Line 1: | Line 1: | ||
| - | [[Image:2ddf.gif|left|200px]]<br /> | + | [[Image:2ddf.gif|left|200px]]<br /><applet load="2ddf" size="350" color="white" frame="true" align="right" spinBox="true" |
| - | <applet load="2ddf" size=" | + | |
caption="2ddf, resolution 1.700Å" /> | caption="2ddf, resolution 1.700Å" /> | ||
'''Crystal structure of TACE in complex with TAPI-2'''<br /> | '''Crystal structure of TACE in complex with TAPI-2'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The crystallization of TNF-alpha converting enzyme (TACE) has been useful | + | The crystallization of TNF-alpha converting enzyme (TACE) has been useful in understanding the structure-activity relationships of new chemical entities. However, the propensity of TACE to undergo autoproteolysis has made enzyme handling difficult and impeded the identification of inhibitor soakable crystal forms. The autoproteolysis of TACE was found to be specific (Y352-V353) and occurred within a flexible loop that is in close proximity to the P-side of the active site. The rate of autoproteolysis was found to be proportional to the concentration of TACE, suggesting a bimolecular reaction mechanism. A limited specificity study of the S(1)' subsite was conducted using surrogate peptides and suggested substitutions that would stabilize the proteolysis of the loop at positions Y352-V353. Two mutant proteases (V353G and V353S) were generated and proved to be highly resistant to autoproteolysis. The kinetics of the more resistant mutant (V353G) and wild-type TACE were compared and demonstrated virtually identical IC(50) values for a panel of competitive inhibitors. However, the k(cat)/K(m) of the mutant for a larger substrate (P6 - P(6)') was approximately 5-fold lower than that for the wild-type enzyme. Comparison of the complexed wild-type and mutant structures indicated a subtle shift in a peripheral P-side loop (comprising the mutation site) that may be involved in substrate binding/turnover and might explain the mild kinetic difference. The characterization of this stabilized form of TACE has yielded an enzyme with similar native kinetic properties and identified a novel crystal form that is suitable for inhibitor soaking and structure determination. |
==About this Structure== | ==About this Structure== | ||
| - | 2DDF is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens] with ZN, CA, INN, IMD, CIT and IPA as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/ADAM_17_endopeptidase ADAM 17 endopeptidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.24.86 3.4.24.86] Full crystallographic information is available from [http:// | + | 2DDF is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens] with <scene name='pdbligand=ZN:'>ZN</scene>, <scene name='pdbligand=CA:'>CA</scene>, <scene name='pdbligand=INN:'>INN</scene>, <scene name='pdbligand=IMD:'>IMD</scene>, <scene name='pdbligand=CIT:'>CIT</scene> and <scene name='pdbligand=IPA:'>IPA</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/ADAM_17_endopeptidase ADAM 17 endopeptidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.24.86 3.4.24.86] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2DDF OCA]. |
==Reference== | ==Reference== | ||
| Line 24: | Line 23: | ||
[[Category: tace adam17 zn-endopeptidase]] | [[Category: tace adam17 zn-endopeptidase]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:57:44 2008'' |
Revision as of 14:57, 21 February 2008
|
Crystal structure of TACE in complex with TAPI-2
Overview
The crystallization of TNF-alpha converting enzyme (TACE) has been useful in understanding the structure-activity relationships of new chemical entities. However, the propensity of TACE to undergo autoproteolysis has made enzyme handling difficult and impeded the identification of inhibitor soakable crystal forms. The autoproteolysis of TACE was found to be specific (Y352-V353) and occurred within a flexible loop that is in close proximity to the P-side of the active site. The rate of autoproteolysis was found to be proportional to the concentration of TACE, suggesting a bimolecular reaction mechanism. A limited specificity study of the S(1)' subsite was conducted using surrogate peptides and suggested substitutions that would stabilize the proteolysis of the loop at positions Y352-V353. Two mutant proteases (V353G and V353S) were generated and proved to be highly resistant to autoproteolysis. The kinetics of the more resistant mutant (V353G) and wild-type TACE were compared and demonstrated virtually identical IC(50) values for a panel of competitive inhibitors. However, the k(cat)/K(m) of the mutant for a larger substrate (P6 - P(6)') was approximately 5-fold lower than that for the wild-type enzyme. Comparison of the complexed wild-type and mutant structures indicated a subtle shift in a peripheral P-side loop (comprising the mutation site) that may be involved in substrate binding/turnover and might explain the mild kinetic difference. The characterization of this stabilized form of TACE has yielded an enzyme with similar native kinetic properties and identified a novel crystal form that is suitable for inhibitor soaking and structure determination.
About this Structure
2DDF is a Single protein structure of sequence from Homo sapiens with , , , , and as ligands. Active as ADAM 17 endopeptidase, with EC number 3.4.24.86 Full crystallographic information is available from OCA.
Reference
Stabilization of the autoproteolysis of TNF-alpha converting enzyme (TACE) results in a novel crystal form suitable for structure-based drug design studies., Ingram RN, Orth P, Strickland CL, Le HV, Madison V, Beyer BM, Protein Eng Des Sel. 2006 Apr;19(4):155-61. Epub 2006 Feb 3. PMID:16459338
Page seeded by OCA on Thu Feb 21 16:57:44 2008
Categories: ADAM 17 endopeptidase | Homo sapiens | Single protein | Orth, P. | CA | CIT | IMD | INN | IPA | ZN | Tace adam17 zn-endopeptidase
