In this study, we focus on the biomimetic development of small molecules and their biological sensing with DNA. The binding of herring sperm deoxyribonucleic acid (hs-DNA) with naturally occurring bioactive small molecule α-amyrin acetate (α-AA), a biomimetic – isolated from the leaves of Ficus (F.) arnottiana is investigated. Collective information from various imaging, spectroscopic and biophysical experiments provides evidence that α-AA is a minor groove sensor of hs-DNA and preferentially binds to the A–T-rich regions. Interactions of different concentrations of small molecule α-AA with hsDNA were evaluated via various analytical techniques such as UV-Vis, circular dichroism (CD) and fluorescence emission spectroscopy. Fluorescence emission spectroscopy results suggest that α-AA decreases the emission level of hsDNA. DNA minor groove sensor Hoechst 33258 and intercalative sensor EB, melting transition analysis (T_M) and viscosity analysis clarified that α-AA binds to hs-DNA via a groove site. Biophysical chemistry and molecular docking studies show that hydrophobic interactions play a major role in this binding. The present research deals with a natural product biosynthesis-linked chemical–biology interface sensor as a biological probe for α-AA: hs-DNA.