Mycobacterium tuberculosis shikimate kinase (MtSK) is a key enzyme involved in the shikimic acid pathway for the biosynthesis of precursors of many important metabolites such as aromatic amino acids, folic acid, and quinones. The presence of this enzyme is essential for the survival of Mycobacterium tuberculosis and absent in mammals. In this study, a pharmacophore-based virtual screening was carried out on 564 secondary metabolite compounds in Aspergillus sp to obtain MtSK inhibitors. This screening initially identified 17 hit compounds, further evaluated based on molecular docking, identifying five natural lead compounds with good binding affinity and interaction. The study identified sydowic acid, tensidol B, pravastatin, lovastatin acid, and yanuthone D potentially inhibit the activity of the desired enzyme and help to fight against tuberculosis, and thus it requires further evaluation by in vitro and in vivo studies. © 2025 Author(s).