*Article* **Potential Efficacy of** *β***-Amyrin Targeting Mycobacterial Universal Stress Protein by In Vitro and In Silico Approach**

**Md Amjad Beg <sup>1</sup> , Shivangi <sup>2</sup> , Obaid Afzal 3,\* , Md Sayeed Akhtar <sup>4</sup> , Abdulmalik S. A. Altamimi <sup>3</sup> , Afzal Hussain <sup>5</sup> , Md Ali Imam <sup>1</sup> , Mohammad Naiyaz Ahmad 6,7 , Sidharth Chopra 6,7 and Fareeda Athar 1,\***


**Abstract:** The emergence of drug resistance and the limited number of approved antitubercular drugs prompted identification and development of new antitubercular compounds to cure *Tuberculosis* (TB). In this work, an attempt was made to identify potential natural compounds that target mycobacterial proteins. Three plant extracts (*A. aspera*, *C. gigantea* and *C. procera*) were investigated. The ethyl acetate fraction of the aerial part of *A. aspera* and the flower ash of *C. gigantea* were found to be effective against *M. tuberculosis* H37Rv. Furthermore, the GC-MS analysis of the plant fractions confirmed the presence of active compounds in the extracts. The *Mycobacterium* target proteins, i.e., available PDB dataset proteins and proteins classified in virulence, detoxification, and adaptation, were investigated. A total of ten target proteins were shortlisted for further study, identified as follows: *BpoC*, *RipA*, *MazF4*, *RipD*, *TB15.3*, *VapC15*, *VapC20*, *VapC21*, *TB31.7*, and *MazF9*. Molecular docking studies showed that *β*-amyrin interacted with most of these proteins and its highest binding affinity was observed with *Mycobacterium* Rv1636 (*TB15.3*) protein. The stability of the protein-ligand complex was assessed by molecular dynamic simulation, which confirmed that *β*-amyrin most firmly interacted with *Rv1636* protein. *Rv1636* is a universal stress protein, which regulates *Mycobacterium* growth in different stress conditions and, thus, targeting *Rv1636* makes *M. tuberculosis* vulnerable to host-derived stress conditions.

**Keywords:** *A. aspera*; *β*-amyrin; *C. gigantea*; *C. procera*; GC-MS; Minimum inhibitory concentration (MIC); Molecular docking; MD simulations; *Tuberculosis* (TB)
