Hydrophobic thermoplastic elastomers, e.g., poly(styrene-b-isobutylene-b-styrene) (SIBS), have found various in vivo biomedical applications. It has long been recognized that biomaterials can be adversely affected by bacterial contamination and clinical infection. However, inhibiting bacterial colonization while simultaneously preserving or enhancing tissue-cell/material interactions is a great challenge. Herein, SIBS substrates were functionalized with nucleases under mild conditions, through polycarboxylate grafts as intermediate. It was demonstrated that the nuclease-modified SIBS could effectively prevent bacterial adhesion and biofilm formation. Cell adhesion assays confirmed that nuclease coatings generally had no negative effects on L929 cell adhesion, compared with the virgin SIBS reference. Therefore, the as-reported nuclease coating may present a promising approach to inhibit bacterial infection, while preserving tissue-cell integration on polymeric biomaterials.