Maria-Solano, Miguel A. and Kinateder, Thomas and Iglesias-Fernandez, Javier and Sterner, Reinhard and Osuna, Silvia (2021) In Silico Identification and Experimental Validation of Distal Activity-Enhancing Mutations in Tryptophan Synthase. ACS CATALYSIS, 11 (21). pp. 13733-13743. ISSN 2155-5435
Full text not available from this repository. (Request a copy)Abstract
Allostery is a central mechanism for the regulation of multi-enzyme complexes. The mechanistic basis that drives allosteric regulation is poorly understood but harbors key information for enzyme engineering. In the present study, we focus on the tryptophan synthase complex that is composed of TrpA and TrpB subunits, which allosterically activate each other. Specifically, we develop a rational approach for identifying key amino acid residues of TrpB distal from the active site. Those residues are predicted to be crucial for shifting the inefficient conformational ensemble of the isolated TrpB to a productive ensemble through intra-subunit allosteric effects. The experimental validation of the conformationally driven TrpB design demonstrates its superior stand-alone activity in the absence of TrpA, comparable to those enhancements obtained after multiple rounds of experimental laboratory evolution. Our work evidences that the current challenge of distal active site prediction for enhanced function in computational enzyme design has become within reach.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | ALLOSTERIC REGULATION; MULTIENZYME COMPLEX; COMPUTATIONAL TOOLS; CRYSTAL-STRUCTURE; EVOLUTION; SUBUNIT; HYPERTHERMOPHILE; ENSEMBLES; TRPB; tryptophan synthase; distal mutations; allostery; enzyme design; shortest path map; ancestral sequence reconstruction |
| Subjects: | 500 Science > 540 Chemistry & allied sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Reinhard Sterner |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 25 Aug 2022 06:17 |
| Last Modified: | 25 Aug 2022 06:17 |
| URI: | https://pred.uni-regensburg.de/id/eprint/46973 |
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