Evaluation of Types of Interactions in Subunit Association in Bacillus subtilis Adenylosuccinate Lyase
| العنوان: | Evaluation of Types of Interactions in Subunit Association in Bacillus subtilis Adenylosuccinate Lyase |
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| المؤلفون: | Lushanti De Zoysa Ariyananda, Roberta F. Colman |
| المصدر: | Biochemistry. 47:2923-2934 |
| بيانات النشر: | American Chemical Society (ACS), 2008. |
| سنة النشر: | 2008 |
| مصطلحات موضوعية: | Models, Molecular, Stereochemistry, Protein subunit, Bacillus subtilis, Biochemistry, Protein Structure, Secondary, Hydrophobic effect, Tetramer, Computer Simulation, Adenylosuccinate lyase, chemistry.chemical_classification, biology, Chemistry, Circular Dichroism, Mutagenesis, Adenylosuccinate Lyase, Temperature, Hydrogen-Ion Concentration, biology.organism_classification, Molecular Weight, Protein Subunits, Enzyme, Mutagenesis, Site-Directed, Electrophoresis, Polyacrylamide Gel, Dimerization, Hydrophobic and Hydrophilic Interactions, Protein Binding, Homotetramer |
| الوصف: | Adenylosuccinate lyase (ASL) of Bacillus subtilis is a homotetramer in which three subunits contribute to each of four active sites. We sought to evaluate the types of interactions responsible for subunit association by studying the enzyme's oligomeric structure at low temperatures as compared to 25 degrees C, in the presence of KBr and after mutagenesis. Analytical ultracentrifugation data reveal that at 25 degrees C ASL is active and exists as 100% tetramer, while at 8 and 4 degrees C, as hydrophobic interactions are weakened, the catalytic activity decreases strikingly and the enzyme dissociates to a mixture of monomer-dimer-trimer, with small amounts of tetramer. In the presence of increasing concentrations of KBr (0.1-2.5 M), which disrupts electrostatic interactions, ASL is dissociated initially to monomer-dimer, with small amounts of trimer-tetramer, and then the monomer species predominates along with small amounts of trimer-tetramer. Very low enzymatic activity was found under these conditions. Accordingly, we postulate that electrostatic interactions are a major source of oligomeric stabilization of B. subtilis ASL. We selected for mutagenesis the closest charged residues (His (299)/Glu (239) and Arg (167)/Asp (217) pairs) located in the subunit interface that has the largest surface area. All of the mutants have low V max values, high K M values, and decreased molecular masses. We conclude that both hydrophobic and electrostatic interactions play roles in maintaining the ASL tetramer and this structure is essential for adenylosuccinate lyase activity. |
| تدمد: | 1520-4995 0006-2960 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::a289b8248dc201c3cc642323a6f9e654 https://doi.org/10.1021/bi701400c |
| رقم الأكسشن: | edsair.doi.dedup.....a289b8248dc201c3cc642323a6f9e654 |
| قاعدة البيانات: | OpenAIRE |
Find this issue from the American Chemical Society | تدمد: | 15204995 00062960 |
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