Less Unfavorable Salt Bridges on the Enzyme Surface Result in More Organic Cosolvent Resistance. Issue 20 (7th April 2021)

• Record Type: Journal Article
• Authors: Cui, Haiyang; Eltoukhy, Lobna; Zhang, Lingling; Markel, Ulrich; Jaeger, Karl‐Erich; Davari, Mehdi D.; Schwaneberg, Ulrich
• Is Part Of: Angewandte Chemie international edition. Volume 60:Issue 20(2021)
• Page Start: 11448
• Page End: 11456
• Publication Date: 2021-04-07
• Subjects: bacillus subtilits lipase A (BSLA) -- directed evolution -- organic solvent resistance -- rational design -- salt bridge; Chemistry -- Periodicals; 540
• DOI: 10.1002/anie.202101642
• Languages: English
• ISSNs: 1433-7851

Abstract

Abstract: Biocatalysis for the synthesis of fine chemicals is highly attractive but usually requires organic (co‐)solvents (OSs). However, native enzymes often have low activity and resistance in OSs and at elevated temperatures. Herein, we report a smart salt bridge design strategy for simultaneously improving OS resistance and thermostability of the model enzyme, Bacillus subtilits Lipase A (BSLA). We combined comprehensive experimental studies of 3450 BSLA variants and molecular dynamics simulations of 36 systems. Iterative recombination of four beneficial substitutions yielded superior resistant variants with up to 7.6‐fold (D64K/D144K) improved resistance toward three OSs while exhibiting significant thermostability (thermal resistance up to 137‐fold, and half‐life up to 3.3‐fold). Molecular dynamics simulations revealed that locally refined flexibility and strengthened hydration jointly govern the highly increased resistance in OSs and at 50–100 °C. The salt bridge redesign provides protein engineers with a powerful and likely general approach to design OSs‐ and/or thermal‐resistant lipases and other α/β‐hydrolases. Abstract : By removing unfavorable surface salt bridges, the organic solvent and thermal resistance of enzymes can be significantly improved. This design strategy results in locally refined flexibility and strengthened hydration.