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B-factor Guided Proline Substitutions in Chromobacterium violaceum Amine Transaminase – An Evaluation of the Proline Rule as a Method for Enzyme Stabilization
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- Land, Henrik (författare)
- Uppsala universitet,KTH,Industriell bioteknologi,Molekylär biomimetik,KTH Royal Inst Technol, Sch Engn Sci Chem Biotechnol & Hlth CBH, Dept Ind Biotechnol, AlbaNova Univ Ctr, S-10691 Stockholm, Sweden,fotosyntesgruppen
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- Campillo-Brocal, Jonatan C. (författare)
- KTH,Industriell bioteknologi,KTH Royal Inst Technol, Sch Engn Sci Chem Biotechnol & Hlth CBH, Dept Ind Biotechnol, AlbaNova Univ Ctr, S-10691 Stockholm, Sweden
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- Svedendahl Humble, Maria (författare)
- Pharem Biotech AB, Biovat Pk,Forskargatan 20 J, S-15136 Sodertalje, Sweden
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- 2019-03-19
- 2019
- Engelska.
- Ingår i: ChemBioChem. - : Wiley-VCH Verlagsgesellschaft. - 1439-4227 .- 1439-7633. ; 20:10, s. 1297-1304
- Tidskriftsartikel (refereegranskat)
Abstract
Ämnesord
Stäng
- Biocatalysis is attracting interest in the chemical industry as a sustainable alternative in large-scale chemical transformations. However, low operational stability of naturally evolved enzymes is a challenge and major efforts are required to engineer protein stability, usually by directed evolution. The development of methods for protein stabilization based on rational design is of great interest, as it would minimize the efforts needed to generate stable enzymes. We hereby present a rational design strategy based on proline substitutions in flexible areas of the protein identified by analyzing B-factors. Several proline substitutions in the amine transaminase from Chromobacterium violaceum were shown to have a positive impact on stability with increased half-life at 60°C by a factor of 2.7 (variant K69P/D218P/K304P/R432P) as well as increased melting temperature by 8.3°C (variant K167P). Finally, the presented method utilizing B-factor analysis in combination with the Proline rule was deemed successful at increasing the stability of this enzyme.
Ämnesord
- NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
Nyckelord
- Biocatalysis
- Enzymes
- Enzyme Stabilization
- Protein engineering
- Molecular modeling
- Bioteknologi
- Biotechnology
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