The invention provides an enzyme-responsive intelligent bacterial adhesion resistant and bactericidal layer-by-layer self-assembled multi-layer film coating and a preparation method thereof. A surface-hydrophobic gel coating which is stable in a water environment is obtained through physical coating of chitosan, hyaluronic acid and a lysine solution, so that a material is endowed with bacterial adhesion resisting and contact sterilizing functions; polylysine improves the bacterial adhesion resistance of the material, so that the coating has the favorable gram-positive and gram-negative bacterial adhesion resisting and sterilizing capabilities. The preparation method is simple in process, quick, mild in condition, and easy for dip-coating, spin-coating, spray-coating and layer-by-layer self-assembly, can be implemented industrially, and has a wide application range, and the antibacterial performance, the bio-compatibility and the lubricity of the surface of medical devices can be effectively improved.