In medical device design, there is a vital need for a coating that promotes treatment of the patient and simultaneously prevents fouling by biomacromolecules which in turn can progress to infections, thrombosis, and other device-related complications. In this work, hydrophobin SC3 (SC3), a self-assembling amphiphilic protein, was coated on a nitric oxide (NO) releasing medical grade polymer to provide an antifouling layer to work synergistically with NO's bactericidal and antiplatelet activity (SC3-NO). The contact angle of SC3 samples were ∼30% lesser than uncoated control samples and was maintained for a month in physiological conditions, demonstrating a stable, hydrophilic coating. NO release characteristics were not adversely affected by the SC3 coating and samples with SC3 coating maintained NO release. Fibrinogen adsorption was reduced over tenfold on SC3 coated samples when compared to non-SC3 coated samples. The viable cell count of adhered bacteria (Staphylococcus aureus) on SC3-NO was 79.097 ± 7.529% lesser than control samples and 49.533 ± 18.18% lesser than NO samples. Platelet adherence on SC3-NO was reduced by 73.407 ± 14.59% when compared to control samples and 53.202 ± 25.67 when compared to NO samples. Finally, the cytocompatibility of SC3-NO was tested and proved to be safe and not trigger a cytotoxic response. The overall favorable results from the physical, chemical and biological characterization analyses demonstrate the novelty and importance of a naturally-produced antifouling layer coated on a bactericidal and antiplatelet polymer, and thus will prove to be advantageous in a multitude of medical device applications.