The development of multi-drug-resistant bacterial infections seriously threatens public health. Hence, efforts are needed to develop a new class of safe and effective antibacterial agents. Two-dimensional molybdenum disulfide (MoS₂)-based nanostructures have great potential as antibacterial agents, but their aggregation limits their further biomedical applications. Here, we report a bio-inspired synthesis of surface-modified MoS₂ nanoflowers (NFs) with a nature motif L-cysteine that show good colloidal stability in aqueous media. The formation of surface-modified MoS₂ NFs has been confirmed using XRD, SEM, TEM, XPS, FTIR, and TGA analysis. The antibacterial activity of as-prepared MoS₂ NFs examined over Gram-negative Escherichia coli (E. coli) and Gram-positive Staphylococcus aureus (S. aureus) bacteria have shown excellent bactericidal activity. The Scanning Electron Microscopy (SEM) images of the bacteria confirm the membrane-directed antibacterial mechanisms where the nanosheets (NSs) in the NFs act as nanoblades and cause cell membrane damage. The antibacterial mechanisms of MoS₂-cys NFs are primarily attributed to membrane damage and the generation of oxidative stresses, which destroy both bacterial strains. The generation of oxidative stress can occur through reactive oxygen species (ROS)-dependent and -independent pathways, as confirmed using flow cytometry and fluorescence imaging and Ellman's assay, respectively. The excessive generation of ROS leads to the inactivation of the bacterial antioxidant defense mechanism. Moreover, the toxicity studies towards human foreskin fibroblast (HFF) cell lines suggested the good biocompatibility of these as-synthesized NFs. We report the intrinsic antibacterial efficiency of MoS₂-cys NFs without any external stimulus (light, H₂O₂, etc.), doping, or drug loading. Our study indicates that the appropriate surface modification of the flower-like morphology of MoS₂ can enhance their colloidal stability and intrinsic antibacterial potency for further applications such as antibacterial coatings, water disinfection, and wound healing.