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| - | [[Image:1m6t.jpg|left|200px]] | + | {{Seed}} |
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| | {{STRUCTURE_1m6t| PDB=1m6t | SCENE= }} | | {{STRUCTURE_1m6t| PDB=1m6t | SCENE= }} |
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| - | '''CRYSTAL STRUCTURE OF B562RIL, A REDESIGNED FOUR HELIX BUNDLE'''
| + | ===CRYSTAL STRUCTURE OF B562RIL, A REDESIGNED FOUR HELIX BUNDLE=== |
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| - | ==Overview==
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| - | To test whether it is practical to use phage display coupled with proteolysis for protein design, we used this approach to convert a partially unfolded four-helix bundle protein, apocytochrome b(562), to a stably folded four-helix bundle protein. Four residues expected to form a hydrophobic core were mutated. One residue was changed to Trp to provide a fluorescence probe for studying the protein's physical properties and to partially fill the void left by the heme. The other three positions were randomly mutated. In addition, another residue in the region to be redesigned was substituted with Arg to provide a specific cutting site for protease Arg-c. This library of mutants was displayed on the surface of phage and challenged with protease Arg-c to select stably folded proteins. The consensus sequence that emerged from the selection included hydrophobic residues at only one of the three positions and non-hydrophobic residues at the other two. Nevertheless, the selected proteins were thermodynamically very stable. The structure of a selected protein was characterized using multi-dimensional NMR. All four helices were formed in the structure. Further, site-directed mutagenesis was used to change one of the two non-hydrophobic residues to a hydrophobic residue, which increased the stability of the protein, indicating that the selection result was not based solely on the protein's global stability and that local structural characteristics may also govern the selection. This conclusion is supported by the crystal structure of another mutant that has two hydrophobic residues substituted for the two non-hydrophobic residues. These results suggest that the hydrophobic interactions in the core are not sufficient to dictate the selection and that the location of the cutting site of the protease also influences the selection of structures.
| + | The line below this paragraph, {{ABSTRACT_PUBMED_12381319}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 12381319 is the PubMed ID number. |
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| | + | {{ABSTRACT_PUBMED_12381319}} |
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| | ==About this Structure== | | ==About this Structure== |
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| | [[Category: Takei, J.]] | | [[Category: Takei, J.]] |
| | [[Category: Protein design]] | | [[Category: Protein design]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sat May 3 00:42:21 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Wed Jul 2 23:20:20 2008'' |
Revision as of 20:20, 2 July 2008
Template:STRUCTURE 1m6t
CRYSTAL STRUCTURE OF B562RIL, A REDESIGNED FOUR HELIX BUNDLE
Template:ABSTRACT PUBMED 12381319
About this Structure
1M6T is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.
Reference
Redesign of a four-helix bundle protein by phage display coupled with proteolysis and structural characterization by NMR and X-ray crystallography., Chu R, Takei J, Knowlton JR, Andrykovitch M, Pei W, Kajava AV, Steinbach PJ, Ji X, Bai Y, J Mol Biol. 2002 Oct 18;323(2):253-62. PMID:12381319
Page seeded by OCA on Wed Jul 2 23:20:20 2008
