Reetz, Manfred T. and Rentzsch, Martin and Pletsch, Andreas and Taglieber, Andreas and Hollmann, Frank and Mondiere, Regis J. G. and Dickmann, Norbert and Hoecker, Birte and Cerrone, Simona and Haeger, Michaela C. and Sterner, Reinhard (2008) A robust protein host for anchoring chelating ligands and organocatalysts. CHEMBIOCHEM, 9 (4). pp. 552-564. ISSN 1439-4227
Full text not available from this repository. (Request a copy)Abstract
In order to put the previously proposed concept of directed evolution of hybrid catalysts (proteins that harbor synthetic transition-metal catalysts or orgonocatalysts) into practice, several prerequisites must be met. The availability of a robust host protein that can be expressed in sufficiently large amounts, and that can be purified in a simple manner is crucial. The thermostable enzyme tHisF from Thermotoga maritima, which constitutes the synthase subunit of a bi-enzyme complex that is instrumental in the biosynthesis of histidine, fulfills these requirements. In the present study, fermentation has been miniaturized and parallelized, as has purification of the protein by simple heat treatment. Several mutants with strategically placed cysteines for subsequent bioconjugation have been produced. One of the tHisF mutants, Cys9Ala/Asp 11Cys, was subjected to bioconjugation by the introduction of a variety of ligands for potential metal ligation, of a ligand/metal moiety, and of several organocatolytic entities that comprise a flavin or thiazolium salts. Characterization by moss spectrometry and tryptic digestion was achieved. As a result of this study, a platform for performing future directed evolution of these hybrid catalysts is now available.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | BAEYER-VILLIGER MONOOXYGENASES; GLYCEROL PHOSPHATE SYNTHASE; DIRECTED EVOLUTION; ARTIFICIAL METALLOENZYMES; ENANTIOSELECTIVE CATALYSIS; HYBRID CATALYSTS; ASYMMETRIC HYDROGENATION; SATURATION MUTAGENESIS; CHEMICAL-MODIFICATION; STRUCTURAL EVIDENCE; bioconjugation; directed evolution; enzymes hybrid catalysts; transition metals |
| Subjects: | 500 Science > 540 Chemistry & allied sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Reinhard Sterner |
| Depositing User: | Petra Gürster |
| Date Deposited: | 14 Jan 2021 12:29 |
| Last Modified: | 14 Jan 2021 12:29 |
| URI: | https://pred.uni-regensburg.de/id/eprint/31190 |
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