دورية أكاديمية
Specific Mutations Reverse Regulatory Effects of Adenosine Phosphates and Increase Their Binding Stoichiometry in CBS Domain-Containing Pyrophosphatase
| العنوان: | Specific Mutations Reverse Regulatory Effects of Adenosine Phosphates and Increase Their Binding Stoichiometry in CBS Domain-Containing Pyrophosphatase |
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| المؤلفون: | Viktor A. Anashkin, Elena A. Kirillova, Victor N. Orlov, Alexander A. Baykov |
| المصدر: | International Journal of Molecular Sciences, Vol 25, Iss 11, p 5768 (2024) |
| بيانات النشر: | MDPI AG, 2024. |
| سنة النشر: | 2024 |
| المجموعة: | LCC:Biology (General) LCC:Chemistry |
| مصطلحات موضوعية: | allosteric regulation, cooperativity, cystathionine β-synthase domain, diadenosine tetraphosphate, enzyme regulation, isothermal calorimetry, Biology (General), QH301-705.5, Chemistry, QD1-999 |
| الوصف: | Regulatory cystathionine β-synthase (CBS) domains are widespread in proteins; however, difficulty in structure determination prevents a comprehensive understanding of the underlying regulation mechanism. Tetrameric microbial inorganic pyrophosphatase containing such domains (CBS-PPase) is allosterically inhibited by AMP and ADP and activated by ATP and cell alarmones diadenosine polyphosphates. Each CBS-PPase subunit contains a pair of CBS domains but binds cooperatively to only one molecule of the mono-adenosine derivatives. We used site-directed mutagenesis of Desulfitobacterium hafniense CBS-PPase to identify the key elements determining the direction of the effect (activation or inhibition) and the “half-of-the-sites” ligand binding stoichiometry. Seven amino acid residues were selected in the CBS1 domain, based on the available X-ray structure of the regulatory domains, and substituted by alanine and other residues. The interaction of 11 CBS-PPase variants with the regulating ligands was characterized by activity measurements and isothermal titration calorimetry. Lys100 replacement reversed the effect of ADP from inhibition to activation, whereas Lys95 and Gly118 replacements made ADP an activator at low concentrations but an inhibitor at high concentrations. Replacement of these residues for alanine increased the stoichiometry of mono-adenosine phosphate binding by twofold. These findings identified several key protein residues and suggested a “two non-interacting pairs of interacting regulatory sites” concept in CBS-PPase regulation. |
| نوع الوثيقة: | article |
| وصف الملف: | electronic resource |
| اللغة: | English |
| تدمد: | 1422-0067 1661-6596 |
| Relation: | https://www.mdpi.com/1422-0067/25/11/5768; https://doaj.org/toc/1661-6596; https://doaj.org/toc/1422-0067 |
| DOI: | 10.3390/ijms25115768 |
| URL الوصول: | https://doaj.org/article/eb9c42e448a34e76b73ada3677b08748 |
| رقم الأكسشن: | edsdoj.b9c42e448a34e76b73ada3677b08748 |
| قاعدة البيانات: | Directory of Open Access Journals |
Full Text Finder | تدمد: | 14220067 16616596 |
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| DOI: | 10.3390/ijms25115768 |