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Engineering of a fe...
Engineering of a femtomolar affinity binding protein to human serum albumin
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- Jonsson, Andreas, 1974- (författare)
- KTH,Molekylär Bioteknologi,Biotherapy and Bacterial Display
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- Dogan, Jakob (författare)
- KTH,Molekylär Bioteknologi
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Harne, Nina (författare)
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Abrahmsén, Lars (författare)
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- Nygren, Per-Åke (författare)
- KTH,Molekylär Bioteknologi
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(creator_code:org_t)
- 2008-05-13
- 2008
- Engelska.
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Ingår i: Protein Engineering Design & Selection. - : Oxford University Press (OUP). - 1741-0126 .- 1741-0134. ; 21:8, s. 515-527
- Relaterad länk:
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https://academic.oup...
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
Ämnesord
Stäng
- We describe the development of a novel serum albumin binding protein showing an extremely high affinity (K(D)) for HSA in the femtomolar range. Using a naturally occurring 46-residue three-helix bundle albumin binding domain (ABD) of nanomolar affinity for HSA as template, 15 residues were targeted for a combinatorial protein engineering strategy to identify variants showing improved HSA affinities. Sequencing of 55 unique phage display-selected clones showed a strong bias for wild-type residues at nine positions, whereas various changes were observed at other positions, including charge shifts. Additionally, a few non-designed substitutions appeared. On the basis of the sequences of 12 variants showing high overall binding affinities and slow dissociation rate kinetics, a set of seven 'second generation' variants were constructed. One variant denoted ABD035 displaying wild-type-like secondary structure content and excellent thermal denaturation/renaturation properties showed an apparent affinity for HSA in the range of 50-500 fM, corresponding to several orders of magnitude improvement compared with the wild-type domain. The ABD035 variant also showed an improved affinity toward serum albumin from a number of other species, and a capture experiment involving human serum indicated that the selectivity for serum albumin had not been compromised from the affinity engineering.
Ämnesord
- NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
Nyckelord
- affinity/combinatorial protein engineering
- Molecular biology
- Molekylärbiologi
Publikations- och innehållstyp
- ref (ämneskategori)
- art (ämneskategori)
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