Engineering of a femtomolar affinity binding protein to human serum albumin
| العنوان: | Engineering of a femtomolar affinity binding protein to human serum albumin |
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| المؤلفون: | Per-Åke Nygren, Andreas Jonsson, Lars Abrahmsén, Nina Herne, Jakob Dogan |
| المصدر: | Protein Engineering Design and Selection. 21:515-527 |
| بيانات النشر: | Oxford University Press (OUP), 2008. |
| سنة النشر: | 2008 |
| مصطلحات موضوعية: | Binding Sites, biology, Binding protein, Serum albumin, Albumin, Bioengineering, Protein Engineering, Human serum albumin, Biochemistry, Protein Structure, Secondary, Recombinant Proteins, Peptide Library, Protein A/G, biology.protein, medicine, Humans, Protein G, Binding site, Carrier Proteins, Molecular Biology, Serum Albumin, Biotechnology, Binding domain, medicine.drug |
| الوصف: | 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. |
| تدمد: | 1741-0134 1741-0126 |
| DOI: | 10.1093/protein/gzn028 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::2f0ebd4ce239cf3b3795854a9c8941bc https://doi.org/10.1093/protein/gzn028 |
| Rights: | OPEN |
| رقم الانضمام: | edsair.doi.dedup.....2f0ebd4ce239cf3b3795854a9c8941bc |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 17410134 17410126 |
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| DOI: | 10.1093/protein/gzn028 |