1yi9
From Proteopedia
(New page: 200px<br /><applet load="1yi9" size="450" color="white" frame="true" align="right" spinBox="true" caption="1yi9, resolution 1.70Å" /> '''Crystal Structure An...) |
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| - | [[Image:1yi9.gif|left|200px]]<br /><applet load="1yi9" size=" | + | [[Image:1yi9.gif|left|200px]]<br /><applet load="1yi9" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1yi9, resolution 1.70Å" /> | caption="1yi9, resolution 1.70Å" /> | ||
'''Crystal Structure Analysis of the oxidized form of the M314I mutant of Peptidylglycine alpha-Hydroxylating Monooxygenase'''<br /> | '''Crystal Structure Analysis of the oxidized form of the M314I mutant of Peptidylglycine alpha-Hydroxylating Monooxygenase'''<br /> | ||
==Overview== | ==Overview== | ||
| - | Many bioactive peptides require amidation of their carboxy terminus to | + | Many bioactive peptides require amidation of their carboxy terminus to exhibit full biological activity. Peptidylglycine alpha-hydroxylating monooxygenase (PHM; EC 1.14.17.3), the enzyme that catalyzes the first of the two steps of this reaction, is composed of two domains, each of which binds one copper atom (CuH and CuM). The CuM site includes Met(314) and two His residues as ligands. Mutation of Met(314) to Ile inactivates PHM, but has only a minimal effect on the EXAFS spectrum of the oxidized enzyme, implying that it contributes only marginally to stabilization of the CuM site. To characterize the role of Met(314) as a CuM ligand, we determined the structure of the Met(314)Ile-PHM mutant. Since the mutant protein failed to crystallize in the conditions of the original wild-type protein, this structure determination required finding a new crystal form. The Met(314)Ile-PHM mutant structure confirms that the mutation does not abolish CuM binding to the enzyme, but causes other structural perturbations that affect the overall stability of the enzyme and the integrity of the CuH site. To eliminate possible effects of crystal contacts, we redetermined the structure of wt-PHM in the Met(314)Ile-PHM crystal form and showed that it does not differ from the structure of wild-type (wt)-PHM in the original crystals. Met(314)Ile-PHM was also shown to be less stable than wt-PHM by differential scanning calorimetry. Both structural and calorimetric studies point to a structural role for the CuM site, in addition to its established catalytic role. |
==About this Structure== | ==About this Structure== | ||
| - | 1YI9 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Rattus_norvegicus Rattus norvegicus] with CU and GOL as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Peptidylglycine_monooxygenase Peptidylglycine monooxygenase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.14.17.3 1.14.17.3] Full crystallographic information is available from [http:// | + | 1YI9 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Rattus_norvegicus Rattus norvegicus] with <scene name='pdbligand=CU:'>CU</scene> and <scene name='pdbligand=GOL:'>GOL</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Peptidylglycine_monooxygenase Peptidylglycine monooxygenase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.14.17.3 1.14.17.3] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1YI9 OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Rattus norvegicus]] | [[Category: Rattus norvegicus]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Amzel, L | + | [[Category: Amzel, L M.]] |
| - | [[Category: Blackburn, N | + | [[Category: Blackburn, N J.]] |
| - | [[Category: Eipper, B | + | [[Category: Eipper, B A.]] |
| - | [[Category: Mains, R | + | [[Category: Mains, R E.]] |
| - | [[Category: Prigge, S | + | [[Category: Prigge, S T.]] |
[[Category: Siebert, X.]] | [[Category: Siebert, X.]] | ||
[[Category: CU]] | [[Category: CU]] | ||
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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 16:05:34 2008'' |
Revision as of 14:05, 21 February 2008
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Crystal Structure Analysis of the oxidized form of the M314I mutant of Peptidylglycine alpha-Hydroxylating Monooxygenase
Overview
Many bioactive peptides require amidation of their carboxy terminus to exhibit full biological activity. Peptidylglycine alpha-hydroxylating monooxygenase (PHM; EC 1.14.17.3), the enzyme that catalyzes the first of the two steps of this reaction, is composed of two domains, each of which binds one copper atom (CuH and CuM). The CuM site includes Met(314) and two His residues as ligands. Mutation of Met(314) to Ile inactivates PHM, but has only a minimal effect on the EXAFS spectrum of the oxidized enzyme, implying that it contributes only marginally to stabilization of the CuM site. To characterize the role of Met(314) as a CuM ligand, we determined the structure of the Met(314)Ile-PHM mutant. Since the mutant protein failed to crystallize in the conditions of the original wild-type protein, this structure determination required finding a new crystal form. The Met(314)Ile-PHM mutant structure confirms that the mutation does not abolish CuM binding to the enzyme, but causes other structural perturbations that affect the overall stability of the enzyme and the integrity of the CuH site. To eliminate possible effects of crystal contacts, we redetermined the structure of wt-PHM in the Met(314)Ile-PHM crystal form and showed that it does not differ from the structure of wild-type (wt)-PHM in the original crystals. Met(314)Ile-PHM was also shown to be less stable than wt-PHM by differential scanning calorimetry. Both structural and calorimetric studies point to a structural role for the CuM site, in addition to its established catalytic role.
About this Structure
1YI9 is a Single protein structure of sequence from Rattus norvegicus with and as ligands. Active as Peptidylglycine monooxygenase, with EC number 1.14.17.3 Full crystallographic information is available from OCA.
Reference
The catalytic copper of peptidylglycine alpha-hydroxylating monooxygenase also plays a critical structural role., Siebert X, Eipper BA, Mains RE, Prigge ST, Blackburn NJ, Amzel LM, Biophys J. 2005 Nov;89(5):3312-9. Epub 2005 Aug 12. PMID:16100265
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