دورية أكاديمية
Arg40 is critical for stability and activity of Adenylosuccinate lyase; a purine salvage enzyme from Leishmania donovani.
| العنوان: | Arg40 is critical for stability and activity of Adenylosuccinate lyase; a purine salvage enzyme from Leishmania donovani. |
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| المؤلفون: | Mochi JA; School of Life Sciences, Central University of Gujarat, Gandhinagar, 382030, Gujarat, India., Jani J; School of Life Sciences, Central University of Gujarat, Gandhinagar, 382030, Gujarat, India., Tak K; School of Life Sciences, Central University of Gujarat, Gandhinagar, 382030, Gujarat, India; Department of Biology, Indian Institute of Sciences Education and Research (IISER), Bhopal, 462 066, Madhya Pradesh, India., Lodhi KK; School of Life Sciences, Central University of Gujarat, Gandhinagar, 382030, Gujarat, India., Pananghat G; Biology Division, Indian Institute of Sciences Education and Research (IISER), Pune, 411008, Maharashtra, India., Pappachan A; School of Life Sciences, Central University of Gujarat, Gandhinagar, 382030, Gujarat, India. Electronic address: anju.p@cug.ac.in. |
| المصدر: | Archives of biochemistry and biophysics [Arch Biochem Biophys] 2024 Jul; Vol. 757, pp. 110040. Date of Electronic Publication: 2024 May 13. |
| نوع المنشور: | Journal Article |
| اللغة: | English |
| بيانات الدورية: | Publisher: Elsevier Country of Publication: United States NLM ID: 0372430 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1096-0384 (Electronic) Linking ISSN: 00039861 NLM ISO Abbreviation: Arch Biochem Biophys Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: <2000- > : San Diego, CA : Elsevier Original Publication: New York, NY : Academic Press |
| مواضيع طبية MeSH: | Adenylosuccinate Lyase*/genetics , Adenylosuccinate Lyase*/chemistry , Adenylosuccinate Lyase*/metabolism , Leishmania donovani*/enzymology , Leishmania donovani*/genetics , Enzyme Stability* , Arginine*/metabolism , Arginine*/chemistry, Purines/metabolism ; Purines/chemistry ; Protozoan Proteins/genetics ; Protozoan Proteins/chemistry ; Protozoan Proteins/metabolism ; Mutagenesis, Site-Directed ; Catalytic Domain ; Molecular Dynamics Simulation |
| مستخلص: | Purine salvage enzymes have been of significant interest in anti-Leishmanial drug development due to the parasite's critical dependence on this pathway for the supply of nucleotides in the absence of a de novo purine synthesis pathway. Adenylosuccinate lyase (ADSL) one of the key enzymes in this pathway is a homo-tetramer, where the active site is formed by residues from three distinct subunits. Analysis of the subunit interfaces of LdADSL, revealed a conserved Arg40 forming critical inter-subunit interactions and also involved in substrate binding. We hypothesized that mutating this residue can affect both the structural stability and activity of the enzyme. In our study, we used biochemical, biophysical, and computational simulation approaches to understand the structural and functional role of Arg40 in LdADSL. We have replaced Arg40 with an Ala and Glu using site directed mutagenesis. The mutant enzymes were similar to wild-type enzyme in secondary structure and subunit association. Thermal shift assays indicated that the mutations affected the protein stability. Both mutants showed decreased specific activities in both forward and reverse directions with significantly weakened affinities towards succinyl-adenosine monophosphate (SAMP). The mutations resulted in changes in C3 loop conformation and D3 domain rotation. Consequently, the orientation of the active site amino acid residues changed resulting in compromised activity and stability. Studies so far have majorly focused on the ADSL active site for designing drugs against it. Our work indicates that an alternative inhibitory mechanism for the enzyme can be designed by targeting the inter-subunit interface. (Copyright © 2024 Elsevier Inc. All rights reserved.) |
| فهرسة مساهمة: | Keywords: Adenylosuccinate lyase; Drug target; Fluorescence spectroscopy; Leishmania; Molecular dynamics simulations; Purine salvage pathway |
| المشرفين على المادة: | EC 4.3.2.2 (Adenylosuccinate Lyase) 94ZLA3W45F (Arginine) 0 (Purines) 0 (Protozoan Proteins) |
| تواريخ الأحداث: | Date Created: 20240516 Date Completed: 20240614 Latest Revision: 20240621 |
| رمز التحديث: | 20240622 |
| DOI: | 10.1016/j.abb.2024.110040 |
| PMID: | 38750922 |
| قاعدة البيانات: | MEDLINE |
Full Text Finder | تدمد: | 1096-0384 |
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| DOI: | 10.1016/j.abb.2024.110040 |