Towards Understanding Directed Evolution: More than Half of All Amino Acid Positions Contribute to Ionic Liquid Resistance of Bacillus subtilis Lipase A. Issue 6 (18th March 2015)

• Record Type: Journal Article
• Authors: Frauenkron‐Machedjou, Victorine Josiane; Fulton, Alexander; Zhu, Leilei; Anker, Carolin; Bocola, Marco; Jaeger, Karl‐Erich; Schwaneberg, Ulrich
• Is Part Of: Chembiochem. Volume 16:Issue 6(2015)
• Page Start: 937
• Page End: 945
• Publication Date: 2015-03-18
• Subjects: Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pharmaceutical chemistry -- Periodicals; 572
• DOI: 10.1002/cbic.201402682
• Languages: English
• ISSNs: 1439-4227

Abstract

<abstract abstract-type="main" xml:lang="en"> <title>Abstract</title> <p>Ionic liquids (ILs) are attractive (co‐)solvents for biocatalysis. However, in high concentration (&gt;10 % IL), enzymes usually show decreased activity. No general principles have been discovered to improve IL resistance of enzymes by protein engineering. We present a systematic study to elucidate general engineering principles by site saturation mutagenesis on the complete gene <italic>bsla</italic>. Screening in presence of four [BMIM]‐based ILs revealed two unexpected lessons on directed evolution: 1) resistance improvement was obtainable at 50–69 % of all amino acid positions, thus explaining the success of small sized random mutant libraries; 2) 6–13 % of substitutions led to improved resistance. Among these, 66–95 % were substitutions by chemically different amino acids (e.g., aromatic to polar/aliphatic/charged amino acids), thus indicating that mutagenesis methods introducing such changes should, at least for lipases like BSLA, be favored to improve IL resistance.</p> </abstract>