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| - | [[Image:2fld.gif|left|200px]] | + | {{Seed}} |
| | + | [[Image:2fld.png|left|200px]] |
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| | {{STRUCTURE_2fld| PDB=2fld | SCENE= }} | | {{STRUCTURE_2fld| PDB=2fld | SCENE= }} |
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| - | '''I-MsoI Re-Designed for Altered DNA Cleavage Specificity'''
| + | ===I-MsoI Re-Designed for Altered DNA Cleavage Specificity=== |
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| - | ==Overview==
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| - | The reprogramming of DNA-binding specificity is an important challenge for computational protein design that tests current understanding of protein-DNA recognition, and has considerable practical relevance for biotechnology and medicine. Here we describe the computational redesign of the cleavage specificity of the intron-encoded homing endonuclease I-MsoI using a physically realistic atomic-level forcefield. Using an in silico screen, we identified single base-pair substitutions predicted to disrupt binding by the wild-type enzyme, and then optimized the identities and conformations of clusters of amino acids around each of these unfavourable substitutions using Monte Carlo sampling. A redesigned enzyme that was predicted to display altered target site specificity, while maintaining wild-type binding affinity, was experimentally characterized. The redesigned enzyme binds and cleaves the redesigned recognition site approximately 10,000 times more effectively than does the wild-type enzyme, with a level of target discrimination comparable to the original endonuclease. Determination of the structure of the redesigned nuclease-recognition site complex by X-ray crystallography confirms the accuracy of the computationally predicted interface. These results suggest that computational protein design methods can have an important role in the creation of novel highly specific endonucleases for gene therapy and other applications. | + | The line below this paragraph, {{ABSTRACT_PUBMED_16738662}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 16738662 is the PubMed ID number. |
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| | + | {{ABSTRACT_PUBMED_16738662}} |
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| | ==About this Structure== | | ==About this Structure== |
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| | [[Category: Dna]] | | [[Category: Dna]] |
| | [[Category: Homing endonuclease]] | | [[Category: Homing endonuclease]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun May 4 04:01:50 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Jul 6 12:06:16 2008'' |
Revision as of 09:06, 6 July 2008
Template:STRUCTURE 2fld
I-MsoI Re-Designed for Altered DNA Cleavage Specificity
Template:ABSTRACT PUBMED 16738662
About this Structure
2FLD is a Single protein structure of sequence from Monomastix sp.. Full crystallographic information is available from OCA.
Reference
Computational redesign of endonuclease DNA binding and cleavage specificity., Ashworth J, Havranek JJ, Duarte CM, Sussman D, Monnat RJ Jr, Stoddard BL, Baker D, Nature. 2006 Jun 1;441(7093):656-9. PMID:16738662
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