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1kyr

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{{STRUCTURE_1kyr| PDB=1kyr | SCENE= }}
{{STRUCTURE_1kyr| PDB=1kyr | SCENE= }}
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'''Crystal Structure of a Cu-bound Green Fluorescent Protein Zn Biosensor'''
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===Crystal Structure of a Cu-bound Green Fluorescent Protein Zn Biosensor===
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==Overview==
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We designed a green fluorescent protein mutant (BFPms1) that preferentially binds Zn(II) (enhancing fluorescence intensity) and Cu(II) (quenching fluorescence) directly to a chromophore ligand that resembles a dipyrrole unit of a porphyrin. Crystallographic structure determination of apo, Zn(II)-bound, and Cu(II)-bound BFPms1 to better than 1.5 A resolution allowed us to refine metal centers without geometric restraints, to calculate experimental standard uncertainty errors for bond lengths and angles, and to model thermal displacement parameters anisotropically. The BFPms1 Zn(II) site (KD = 50 muM) displays distorted trigonal bipyrimidal geometry, with Zn(II) binding to Glu222, to a water molecule, and tridentate to the chromophore ligand. In contrast, the BFPms1 Cu(II) site (KD = 24 muM) exhibits square planar geometry similar to metalated porphyrins, with Cu(II) binding to the chromophore chelate and Glu222. The apo structure reveals a large electropositive region near the designed metal insertion channel, suggesting a basis for the measured metal cation binding kinetics. The preorganized tridentate ligand is accommodated in both coordination geometries by a 0.4 A difference between the Zn and Cu positions and by distinct rearrangements of Glu222. The highly accurate metal ligand bond lengths reveal different protonation states for the same oxygen bound to Zn vs Cu, with implications for the observed metal ion specificity. Crystallographic anisotropic thermal factor analysis validates metal ion rigidification of the chromophore in enhancement of fluorescence intensity upon Zn(II) binding. Thus, our high-resolution structures reveal how structure-based design has effectively linked selective metal binding to changes in fluorescent properties. Furthermore, this protein Zn(II) biosensor provides a prototype suitable for further optimization by directed evolution to generate metalloprotein variants with desirable physical or biochemical properties.
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{{ABSTRACT_PUBMED_11929238}}
==About this Structure==
==About this Structure==
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[[Category: Chromophore]]
[[Category: Chromophore]]
[[Category: Cu binding]]
[[Category: Cu binding]]
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Wed Jul 2 11:18:41 2008''

Revision as of 08:18, 2 July 2008

Image:1kyr.png

Template:STRUCTURE 1kyr

Crystal Structure of a Cu-bound Green Fluorescent Protein Zn Biosensor

Template:ABSTRACT PUBMED 11929238

About this Structure

1KYR is a Single protein structure of sequence from Aequorea victoria. Full crystallographic information is available from OCA.

Reference

Structural chemistry of a green fluorescent protein Zn biosensor., Barondeau DP, Kassmann CJ, Tainer JA, Getzoff ED, J Am Chem Soc. 2002 Apr 10;124(14):3522-4. PMID:11929238

Page seeded by OCA on Wed Jul 2 11:18:41 2008

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