Preparation of the gold core and silver shell NP (AuAgNP) is challenging because of the facile oxidation of silver. Here such a NP is carefully synthesized and conjugated with subtilisin to arrive at a stable spherical material of 120–130 nm in diameter (AuAgSNP). A biomaterial prepared by immobilizing AuAgSNP on polycaprolactam (PCL) exhibits antibiofilm properties against S. aureus and E. coli, but with lesser potency than the one prepared with bare AuAgNP. Subtilisin degrades the adhesive surface proteins of the bacteria thereby preventing the biofilm formation. Subtilisin conjugated AuAgSNP is not cytotoxic to 3T3 cells at its MIC, in contrast to AuAgNP. The presence of subtilisin promotes the fibroblast proliferation. This study indicates that AuAgSNP has antibacterial/antibiofilm activities as well as biocompatibility unlike NPs which are very cytotoxic to cells. Hence AuAgSNP can be used in medical implants and devices. © 2018