Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches
<i>Mycobacterium tuberculosis</i> has seen tremendous success as it has developed defenses to reside in host alveoli despite various host-related stress circumstances. Rv1636 is a universal stress protein contributing to mycobacterial survival in different host-derived stress conditions....
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MDPI AG
2022-08-01
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| author | Md Amjad Beg Sadaf Anas Shamsi Sibasis Sahoo Mohd Yousuf Mohammad Zeeshan Najm Yahya Ahmad Almutawif Asimul Islam Abdulaziz A. Aloliqi Fareeda Athar |
| author_facet | Md Amjad Beg Sadaf Anas Shamsi Sibasis Sahoo Mohd Yousuf Mohammad Zeeshan Najm Yahya Ahmad Almutawif Asimul Islam Abdulaziz A. Aloliqi Fareeda Athar |
| author_sort | Md Amjad Beg |
| collection | DOAJ |
| description | <i>Mycobacterium tuberculosis</i> has seen tremendous success as it has developed defenses to reside in host alveoli despite various host-related stress circumstances. Rv1636 is a universal stress protein contributing to mycobacterial survival in different host-derived stress conditions. Both ATP and cAMP can be bound with the Rv1636, and their binding actions are independent of one another. β-Amyrin, a triterpenoid compound, is abundant in medicinal plants and has many pharmacological properties and broad therapeutic potential. The current study uses biochemical, biophysical, and computational methods to define the binding of Rv1636 with β-Amyrin. A substantial interaction between β-Amyrin and Rv1636 was discovered by molecular docking studies, which helped decipher the critical residues involved in the binding process. VAL60 is a crucial residue found in the complexes of both Rv1636_β-Amyrin and Rv1636-ATP. Additionally, the Rv1636_β-Amyrin complex was shown to be stable by molecular dynamics simulation studies (MD), with minimal changes observed during the simulation. In silico observations were further complemented by in vitro assays. Successful cloning, expression, and purification of Rv1636 were accomplished using Ni-NTA affinity chromatography. The results of the ATPase activity assay indicated that Rv1636’s ATPase activity was inhibited in the presence of various β-Amyrin concentrations. Additionally, circular dichroism spectroscopy (CD) was used to examine modifications to Rv1636 secondary structure upon binding of β-Amyrin. Finally, isothermal titration calorimetry (ITC) advocated spontaneous binding of β-Amyrin with Rv1636 elucidating the thermodynamics of the Rv1636_β-Amyrin complex. Thus, the study establishes that β-Amyrin binds to Rv1636 with a significant affinity forming a stable complex and inhibiting its ATPase activity. The present study suggests that β-Amyrin might affect the functioning of Rv1636, which makes the bacterium vulnerable to different stress conditions. |
| first_indexed | 2024-03-09T11:52:16Z |
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| language | English |
| last_indexed | 2024-03-09T11:52:16Z |
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| spelling | doaj.art-381a0e373b624cb18af6f126283e3df92023-11-30T23:13:37ZengMDPI AGBiology2079-77372022-08-01118121410.3390/biology11081214Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro ApproachesMd Amjad Beg0Sadaf1Anas Shamsi2Sibasis Sahoo3Mohd Yousuf4Mohammad Zeeshan Najm5Yahya Ahmad Almutawif6Asimul Islam7Abdulaziz A. Aloliqi8Fareeda Athar9Centre for Interdisciplinary Research in Basic Science, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, IndiaDepartment of Biotechnology, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, IndiaCentre for Interdisciplinary Research in Basic Science, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, IndiaMembrane Protein Biology, International Centre for Genetic Engineering and Biotechnology (ICGEB), New Delhi 110067, IndiaCentre for Interdisciplinary Research in Basic Science, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, IndiaSchool of Biosciences, Apeejay Stya University, Gurugram 122103, IndiaDepartment of Medical Laboratories Technology, College of Applied Medical Sciences, Taibah University, Madinah 42353, Saudi ArabiaCentre for Interdisciplinary Research in Basic Science, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, IndiaDepartment of Medical Biotechnology, College of Applied Medical Sciences, Qassim University, Buraydah 51542, Saudi ArabiaCentre for Interdisciplinary Research in Basic Science, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India<i>Mycobacterium tuberculosis</i> has seen tremendous success as it has developed defenses to reside in host alveoli despite various host-related stress circumstances. Rv1636 is a universal stress protein contributing to mycobacterial survival in different host-derived stress conditions. Both ATP and cAMP can be bound with the Rv1636, and their binding actions are independent of one another. β-Amyrin, a triterpenoid compound, is abundant in medicinal plants and has many pharmacological properties and broad therapeutic potential. The current study uses biochemical, biophysical, and computational methods to define the binding of Rv1636 with β-Amyrin. A substantial interaction between β-Amyrin and Rv1636 was discovered by molecular docking studies, which helped decipher the critical residues involved in the binding process. VAL60 is a crucial residue found in the complexes of both Rv1636_β-Amyrin and Rv1636-ATP. Additionally, the Rv1636_β-Amyrin complex was shown to be stable by molecular dynamics simulation studies (MD), with minimal changes observed during the simulation. In silico observations were further complemented by in vitro assays. Successful cloning, expression, and purification of Rv1636 were accomplished using Ni-NTA affinity chromatography. The results of the ATPase activity assay indicated that Rv1636’s ATPase activity was inhibited in the presence of various β-Amyrin concentrations. Additionally, circular dichroism spectroscopy (CD) was used to examine modifications to Rv1636 secondary structure upon binding of β-Amyrin. Finally, isothermal titration calorimetry (ITC) advocated spontaneous binding of β-Amyrin with Rv1636 elucidating the thermodynamics of the Rv1636_β-Amyrin complex. Thus, the study establishes that β-Amyrin binds to Rv1636 with a significant affinity forming a stable complex and inhibiting its ATPase activity. The present study suggests that β-Amyrin might affect the functioning of Rv1636, which makes the bacterium vulnerable to different stress conditions.https://www.mdpi.com/2079-7737/11/8/1214Rv1636β-Amyrinmolecular dockingmolecular dynamics simulationcircular dichroism spectroscopyisothermal titration calorimetry |
| spellingShingle | Md Amjad Beg Sadaf Anas Shamsi Sibasis Sahoo Mohd Yousuf Mohammad Zeeshan Najm Yahya Ahmad Almutawif Asimul Islam Abdulaziz A. Aloliqi Fareeda Athar Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches Biology Rv1636 β-Amyrin molecular docking molecular dynamics simulation circular dichroism spectroscopy isothermal titration calorimetry |
| title | Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches |
| title_full | Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches |
| title_fullStr | Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches |
| title_full_unstemmed | Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches |
| title_short | Mechanistic Insight into the Enzymatic Inhibition of β-Amyrin against Mycobacterial Rv1636: In Silico and In Vitro Approaches |
| title_sort | mechanistic insight into the enzymatic inhibition of β amyrin against mycobacterial rv1636 in silico and in vitro approaches |
| topic | Rv1636 β-Amyrin molecular docking molecular dynamics simulation circular dichroism spectroscopy isothermal titration calorimetry |
| url | https://www.mdpi.com/2079-7737/11/8/1214 |
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