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Conformational stab...
Conformational stabilization of an engineered binding protein.
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- Wahlberg, Elisabet (författare)
- KTH,Skolan för bioteknologi (BIO)
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- Härd, Torleif, 1959 (författare)
- Gothenburg University,Göteborgs universitet,Institutionen för biomedicin, avdelningen för medicinsk kemi och cellbiologi,Institute of Biomedicine, Department of Medical Biochemistry and Cell Biology
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(creator_code:org_t)
- 2006-05-12
- 2006
- Engelska.
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 128:23, s. 7651-60
- Relaterad länk:
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http://pubs.acs.org/...
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https://gup.ub.gu.se...
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https://doi.org/10.1...
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https://urn.kb.se/re...
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Abstract
Ämnesord
Stäng
- We analyzed the thermodynamic basis for improvement of a binding protein by disulfide engineering. The Z(SPA)(-)(1) affibody binds to its Z domain binding partner with a dissociation constant K(d) = 1.6 microM, and previous analyses suggested that the moderate affinity is due to the conformational heterogeneity of free Z(SPA)(-)(1) rather than to a suboptimal binding interface. Studies of five stabilized Z(SPA)(-)(1) double cystein mutants show that it is possible to improve the affinity by an order of magnitude to K(d) = 130 nM, which is close to the range (20 to 70 nM) observed with natural Z domain binders, without altering the protein-protein interface obtained by phage display. Analysis of the binding thermodynamics reveals a balance between conformational entropy and desolvation entropy: the expected and favorable reduction of conformational entropy in the best-binding Z(SPA)(-)(1) mutant is completely compensated by an unfavorable loss of desolvation entropy. This is consistent with a restriction of possible conformations in the disulfide-containing mutant and a reduction of average water-exposed nonpolar surface area in the free state, resulting in a smaller conformational entropy penalty, but also a smaller change in surface area, for binding of mutant compared to wild-type Z(SPA)(-)(1). Instead, higher Z domain binding affinity in a group of eight Z(SPA)(-)(1) variants correlates with more favorable binding enthalpy and enthalpy-entropy compensation. These results suggest that protein-protein binding affinity can be improved by stabilizing conformations in which enthalpic effects can be fully explored.
Ämnesord
- TEKNIK OCH TEKNOLOGIER -- Industriell bioteknik -- Annan industriell bioteknik (hsv//swe)
- ENGINEERING AND TECHNOLOGY -- Industrial Biotechnology -- Other Industrial Biotechnology (hsv//eng)
Nyckelord
- Algorithms
- Calorimetry
- Circular Dichroism
- Cysteine
- chemistry
- genetics
- metabolism
- Disulfides
- chemistry
- Mutation
- Protein Binding
- Protein Conformation
- Protein Denaturation
- Protein Engineering
- Protein Folding
- Proteins
- chemistry
- genetics
- metabolism
- Thermodynamics
- disulfide bonds
- Structural biochemistry
Publikations- och innehållstyp
- ref (ämneskategori)
- art (ämneskategori)
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