Abstract Fusarium oxysporum is one of the most common soil borne pathogens over the world. This work tried to introduce safe and efficient agent in controlling this pathogen infection in cucumber plants, beside that understanding the mechanism at which fabitine enables the cucumber plants to fight wilt disease. Also, this works introduce a molecular modeling for the direct interaction between fabatin and the Fusarium oxysporum. We evaluated the impact of defensin isolated from broad bean debris (fabatin) against the Fusarium wilt disease in the greenhouse, followed by the analysis of defense-related enzymes, including superoxide dismutase, polyphenol oxidase, soluble peroxidase, cell wall-bound peroxidase, catalase, lactate dehydrogenase, phenylalanine ammonia-lyase, chitinase, and glucanase. Moreover, the growth-regulating protein of the fungus (Snt2, a BAH/PHD) was docked with the fabatin. Fabatin reduced the disease severity from 18.52 to 0.0% by 100% disease reduction, 30 days after transplanting, and from 66.67 to 33.33% by 50.01% disease reduction after 60 days. Interestingly fabatin spray induced upregulation of the studied defense enzymes. The sprayed group showed significant continuous increase in defense related enzymes under infection. Furthermore, fabatin showed a high binding affinity to the Bromo-adjacent domain (BAH) of the Snt2 by - 214.42 kacl/mol and this can prevent the transcriptional regulation processes of the fungus. The present study proved that the exogenous spray fabatin is an effective way in controlling cucumber Fusarium wilt. This is achieved through the enhancement the defense related enzymes which regulate the resistance mechanism in the plants. Moreover, fabatin can directly influence the fungal growth.