3CLpro is a viable target for developing antiviral therapies against the coronavirus. With the urgent need to find new possible inhibitors, a structure-based virtual screening approach was developed. This study recognized 75 pharmacologically bioactive compounds from our in-house library of 1052 natural product-based compounds that satisfied drug-likeness criteria and exhibited good bioavailability and membrane permeability. Among these compounds, three promising sulfonamide chalcones were identified by combined theoretical and experimental approaches, with SWC423 being the most suitable representative compound due to its competitive inhibition and low cytotoxicity in Vero E6 cells (EC50 = 0.89 ± 0.32 μM; CC50 = 25.54 ± 1.38 μM; SI = 28.70). The binding and stability of SWC423 in the 3CLpro active site were investigated through all-atom molecular dynamics simulation and fragment molecular orbital calculation, indicating its potential as a 3CLpro inhibitor for further SARS-CoV-2 therapeutic research. These findings suggested that inhibiting 3CLpro with a sulfonamide chalcone such as SWC423 may pave the effective way for developing COVID-19 treatments.