2ia8
From Proteopedia
(New page: 200px<br /><applet load="2ia8" size="350" color="white" frame="true" align="right" spinBox="true" caption="2ia8, resolution 1.48Å" /> '''Kinetic and Crystall...) |
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==Overview== | ==Overview== | ||
| - | Manganese peroxidase (MnP) from the white rot fungus Phanerochaete | + | Manganese peroxidase (MnP) from the white rot fungus Phanerochaete chrysosporium contains a manganese-binding site that plays a critical role in its function. Previously, a Mn(II)-binding site was designed into cytochrome c peroxidase (CcP) based on sequence homology (Yeung et al. in Chem. Biol. 4:215-222, 1997; Gengenbach et al. in Biochemistry 38:11425-11432, 1999). Here, we report a redesign of this site based on X-ray structural comparison of MnP and CcP. The variant, CcP(D37E, V45E, H181E), displays 2.5-fold higher catalytic efficiency (k (cat)/K (M)) than the variant in the original design, mostly due to a stronger K (M) of 1.9 mM (vs. 4.1 mM). High-resolution X-ray crystal structures of a metal-free form and a form with Co(II) at the designed Mn(II) site were also obtained. The metal ion in the engineered metal-binding site overlays well with Mn(II) bound in MnP, suggesting that this variant is the closest structural model of the Mn(II)-binding site in MnP for which a crystal structure exists. A major difference arises in the distances of the ligands to the metal; the metal-ligand interactions in the CcP variant are much weaker than the corresponding interactions in MnP, probably owing to partial occupancy of metal ion at the designed site, difference in the identity of metal ions (Co(II) rather than Mn(II)) and other interactions in the second coordination sphere. These results indicate that the metal ion, the ligands, and the environment around the metal-binding site play important roles in tuning the structure and function of metalloenzymes. |
==About this Structure== | ==About this Structure== | ||
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[[Category: Saccharomyces cerevisiae]] | [[Category: Saccharomyces cerevisiae]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Conaster, C | + | [[Category: Conaster, C D.N.]] |
| - | [[Category: Gao, Y | + | [[Category: Gao, Y G.]] |
| - | [[Category: Gengenbach, A | + | [[Category: Gengenbach, A J.]] |
[[Category: Lu, Y.]] | [[Category: Lu, Y.]] | ||
| - | [[Category: Mirarefi, A | + | [[Category: Mirarefi, A Y.]] |
[[Category: Pfister, T.]] | [[Category: Pfister, T.]] | ||
[[Category: Robinson, H.]] | [[Category: Robinson, H.]] | ||
| - | [[Category: Wang, A | + | [[Category: Wang, A H.J.]] |
[[Category: Zhao, X.]] | [[Category: Zhao, X.]] | ||
| - | [[Category: Zukoski, C | + | [[Category: Zukoski, C F.]] |
[[Category: HEM]] | [[Category: HEM]] | ||
[[Category: biomimetic]] | [[Category: biomimetic]] | ||
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[[Category: protein engineering]] | [[Category: protein engineering]] | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 17:50:32 2008'' |
Revision as of 15:50, 21 February 2008
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Kinetic and Crystallographic Studies of a Redesigned Manganese-Binding Site in Cytochrome c Peroxidase
Overview
Manganese peroxidase (MnP) from the white rot fungus Phanerochaete chrysosporium contains a manganese-binding site that plays a critical role in its function. Previously, a Mn(II)-binding site was designed into cytochrome c peroxidase (CcP) based on sequence homology (Yeung et al. in Chem. Biol. 4:215-222, 1997; Gengenbach et al. in Biochemistry 38:11425-11432, 1999). Here, we report a redesign of this site based on X-ray structural comparison of MnP and CcP. The variant, CcP(D37E, V45E, H181E), displays 2.5-fold higher catalytic efficiency (k (cat)/K (M)) than the variant in the original design, mostly due to a stronger K (M) of 1.9 mM (vs. 4.1 mM). High-resolution X-ray crystal structures of a metal-free form and a form with Co(II) at the designed Mn(II) site were also obtained. The metal ion in the engineered metal-binding site overlays well with Mn(II) bound in MnP, suggesting that this variant is the closest structural model of the Mn(II)-binding site in MnP for which a crystal structure exists. A major difference arises in the distances of the ligands to the metal; the metal-ligand interactions in the CcP variant are much weaker than the corresponding interactions in MnP, probably owing to partial occupancy of metal ion at the designed site, difference in the identity of metal ions (Co(II) rather than Mn(II)) and other interactions in the second coordination sphere. These results indicate that the metal ion, the ligands, and the environment around the metal-binding site play important roles in tuning the structure and function of metalloenzymes.
About this Structure
2IA8 is a Single protein structure of sequence from Saccharomyces cerevisiae with as ligand. Active as Cytochrome-c peroxidase, with EC number 1.11.1.5 Full crystallographic information is available from OCA.
Reference
Kinetic and crystallographic studies of a redesigned manganese-binding site in cytochrome c peroxidase., Pfister TD, Mirarefi AY, Gengenbach AJ, Zhao X, Danstrom C, Conatser N, Gao YG, Robinson H, Zukoski CF, Wang AH, Lu Y, J Biol Inorg Chem. 2007 Jan;12(1):126-37. Epub 2006 Oct 5. PMID:17021923
Page seeded by OCA on Thu Feb 21 17:50:32 2008
Categories: Cytochrome-c peroxidase | Saccharomyces cerevisiae | Single protein | Conaster, C D.N. | Gao, Y G. | Gengenbach, A J. | Lu, Y. | Mirarefi, A Y. | Pfister, T. | Robinson, H. | Wang, A H.J. | Zhao, X. | Zukoski, C F. | HEM | Biomimetic | Manganese oxidation | Metal-binding site | Metalloprotein | Oxidoreductase | Protein engineering
