Kneuttinger, Andrea C. and Rajendran, Chitra and Simeth, Nadja A. and Bruckmann, Astrid and Koenig, Burkhard and Sterner, Reinhard (2020) Significance of the Protein Interface Configuration for Allostery in Imidazole Glycerol Phosphate Synthase. BIOCHEMISTRY, 59 (29). pp. 2729-2742. ISSN 0006-2960,
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Imidazole glycerol phosphate synthase (ImGPS) from Thermotoga maritima is a model enzyme for studying allostery. The ImGPS complex consists of the cyclase subunit HisF and the glutaminase subunit HisH whose activity is stimulated by substrate binding to HisF in a V-type manner. To investigate the significance of a putative closing hinge motion at the cyclase:glutaminase interface for HisH activity, we replaced residue W123 in HisH with the light-switchable unnatural amino acid phenylalanine-4'-azobenzene (AzoF). Crystal structure analysis employing angle, buried surface area, and distance measurements showed that incorporation of AzoF at this position causes a closing of the interface by similar to 18 +/- 3%. This slightly different interface configuration results in a much higher catalytic efficiency in unstimulated HisH due to an elevated turnover number. Moreover, the catalytic efficiency of HisH when stimulated by binding of a substrate to HisF was also significantly increased by AzoF incorporation. This was caused by a K-type stimulation that led to a decrease in the apparent dissociation constant for its substrate, glutamine. In addition, AzoF improved the apparent binding of a substrate analogue at the HisF active site. Remarkably, light-induced isomerization of AzoF considerably enhanced these effects. In conclusion, our findings confirm that signal transduction from HisF to HisH in ImGPS involves the closing of the cyclase:glutaminase subunit interface and that incorporation of AzoF at a hinge position reinforces this catalytically relevant conformational change.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | STRUCTURAL EVIDENCE; BARREL; ISOMERIZATION; EVOLUTION; BIENZYME; CONTACTS; SYSTEMS; |
| Subjects: | 500 Science > 540 Chemistry & allied sciences 500 Science > 570 Life sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Reinhard Sterner Biology, Preclinical Medicine > Institut für Biochemie, Genetik und Mikrobiologie Chemistry and Pharmacy > Institut für Organische Chemie > Lehrstuhl Prof. Dr. Burkhard König |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 17 Mar 2021 15:39 |
| Last Modified: | 17 Mar 2021 15:39 |
| URI: | https://pred.uni-regensburg.de/id/eprint/44150 |
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