## 4 Antibofilm Coatings Based on Antimicrobial Enzymes  
Antimicrobial enzymes emerged as a very attractive bioinspired antibofilm strategy and as a very promising alternative to the classical antibiotic approach since these enzymes widespread among living things act as a defense mechanism against infection from microorganisms being able to directly attack bacteria, interfere with biofilm formation, and/or catalyze reactions that generate antimicrobial compounds. Many enzymes from different classes exhibit antimicrobial activity [21]:  
1. Proteolytic enzymes - Subtilisins that hydrolyze adhesins thereby impeding bacterial attachment onto solid supports; - Lysostaphin, which cleaves the pentaglycine bridges of _Staphylococci_ cell; - Bacteriophage lysins, which are peptidoglycan hydrolases that cause osmotic lysis and cell death of the bacterium;
2. Polysaccharide degrading enzymes - Lysozyme, which catalyzes the hydrolysis of the glycosidic bond between \(N\)-acetylmuramic acid and \(N\)-acetyl-\(D\)-glucosamine residues in peptidoglycan thereby damaging the bacterial cell wall; - Alpha-amylases; - Dispersin B (DspB), which disrupts biofilm formation by catalyzing the hydrolysis of the \(\beta\)-1,6-glycosidic linkages within poly-\(\beta\)-1,6-N-acetyl-D-glucosamine, which is an important polymer needed for attachment of _E. coli_ and _Staphylococcus epidermidis_ biofilms onto surfaces. - Alginate lyase;
3. Oxidative enzymes - The hydrogen peroxide producing enzymes like glucose oxidase (GOx) and cellobiose dehydrogenase (CDH); cellobiose is a reducing disaccharide, which donates electrons getting oxidized to cellobiono-\(\delta\)-lactone, while electrons are accepted by oxygen, which in turn is reduced to hydrogen peroxide (see the redox half-reaction in Scheme 7); the known disinfectant and antiseptic properties of hydrogen peroxide rely on its capacity to generate free radicals able to attack lipid membranes, DNA, and other vital constituents of the bacterial cell; - Hydrogen peroxide using enzymes like haloperoxidases, lactoperoxidase, and myeloperoxidase, which catalyze the oxidation of halide and thiocyanate anions to more potent antimicrobial agents consuming the hydrogen peroxide produced by the hydrogen peroxide producing enzymes;
4. Quorum-quenching enzymes like lactonases, which catalyze the hydrolysis of the ester bond of acylated homoserine lactones thereby impeding the binding of these autoinducers to their target transcriptional regulators;
5. DNases that cleave DNA by catalyzing the hydrolysis of the phosphodieser bonds of the sugar-phosphate backbone; e-DNA derived from lysed bacteria play an important role in the initial establishment of the biofilm and subsequently in the maintenance of the biofilm's structural integrity acting as an effective crosslinker. CDH was covalently immobilized on PDMS following a four-step synthetic pathway developed by [PERSON] et al. (2018). First, PDMS urinary catheters were washed with 96% ethanol and distilled water, and next were subjected to a low-pressure oxygen plasma treatment in order to obtain hydroxy-functionalized PDMS surface. The second step of the functionalization process consisted of the silanization with (3-aminopropyl)-triethoxysilane (APTES) to yield an amine-modified PDMS surface. In the third step, the homobifunctional linker glutaraldehyde (GA) was tethered to the amine-terminated PDMS surface through one of the terminal aldehyde functions. The aldehyde group left free at the other end of the linker was used to covalently attach the enzyme in the fourth step of the synthesis. The surface immobilization did not affect the activity of CDH, as it was demonstrated by measuring the amount of hydrogen peroxide produced by the immobilized enzyme. The enzymatic activity was determined in buffered media of different pH values (5.5; 6.0; 7.3; 8.0). The best enzymatic activity was obtained at pH = 8.0.