Minimalist proteins: Design of new molecular recognition scaffolds
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| 024 | 7 | 0 | |a 10.1351/pac200476071579 |2 doi |
| 035 | |a (NATIONALLICENCE)gruyter-10.1351/pac200476071579 | ||
| 100 | 1 | |a Shin |D J. A. |u Department of Chemistry, University of Toronto, Mississauga, Ontario L5L 1C6, Canada | |
| 245 | 1 | 0 | |a Minimalist proteins: Design of new molecular recognition scaffolds |h [Elektronische Daten] |c [J. A. Shin] |
| 520 | 3 | |a We hypothesize that we can exploit what Nature has already evolved by manipulating the alpha-helix molecular recognition scaffold. Therefore, minimalist proteins capable of sequence-specific, high-affinity binding of DNA were generated to probe how proteins are used and can be used to recognize DNA. The already minimal basic region/leucine zipper motif (bZIP) of GCN4 was reduced to an even more simplified structure by substitution with alanine residues —hence, a generic, Ala-based, helical scaffold. The proteins generated, wt bZIP, 4A,11A, and 18A, contain 0, 4, 11, and 18 alanine mutations in their DNA-binding basic regions, respectively. All alanine mutants still retain alpha-helical structure and DNA-bind- ing function, despite loss of virtually all Coulombic protein-DNA interactions. Mass spectrometry allowed characterization of proteins and post-translational modifications. Fluorescence anisotropy and DNase I footprinting were used to measure in situ binding of these mutant proteins to DNA duplexes containing target sites AP-1 (5'-TGACTCA-3'), ATF/CREB (5'-TGACGTCA-3'),or nonspecific DNA. The roles of van der Waals and Coulombic interactions toward binding specificity and affinity are being investigated. Thus, both DNA-binding specificity and affinity are maintained in all our bZIP derivatives. This Ala-rich scaffold may be useful in design and synthesis of small, alpha-helical proteins with desired DNA-recognition properties. | |
| 540 | |a © 2013 Walter de Gruyter GmbH, Berlin/Boston | ||
| 773 | 0 | |t Pure and Applied Chemistry |d De Gruyter |g 76/7-8(2004-01-01), 1579-1590 |x 0033-4545 |q 76:7-8<1579 |1 2004 |2 76 |o pac | |
| 856 | 4 | 0 | |u https://doi.org/10.1351/pac200476071579 |q text/html |z Onlinezugriff via DOI |
| 908 | |D 1 |a research article |2 jats | ||
| 950 | |B NATIONALLICENCE |P 856 |E 40 |u https://doi.org/10.1351/pac200476071579 |q text/html |z Onlinezugriff via DOI | ||
| 950 | |B NATIONALLICENCE |P 100 |E 1- |a Shin |D J. A. |u Department of Chemistry, University of Toronto, Mississauga, Ontario L5L 1C6, Canada | ||
| 950 | |B NATIONALLICENCE |P 773 |E 0- |t Pure and Applied Chemistry |d De Gruyter |g 76/7-8(2004-01-01), 1579-1590 |x 0033-4545 |q 76:7-8<1579 |1 2004 |2 76 |o pac | ||
| 900 | 7 | |b CC0 |u http://creativecommons.org/publicdomain/zero/1.0 |2 nationallicence | |
| 898 | |a BK010053 |b XK010053 |c XK010000 | ||
| 949 | |B NATIONALLICENCE |F NATIONALLICENCE |b NL-gruyter | ||