Title
Bactericidal Hydrogels via Surface Functionalization with Cecropin A
Author
Megan A Cole, Timothy F Scott, Charlene M. Mello
Year
2016
Journal
ACS Biomaterials Science and Engineering
Abstract
The immobilization of antimicrobial peptides (AMPs) to surfaces, enabling their utilization in biosensor and antibacterial/antifouling coating applications, is typically performed using rigid, solid support materials such as glass or gold and may require lengthy, temperamental protocols. Here, we employ a hydrogel immobilization platform to afford facile fabrication and surface functionalization while offering improved biocompatibility for evaluating the influence of linker length, surface density, and AMP conjugation site on retained peptide activity. Rapid, interfacial photopolymerization using the radical-mediated thiol–ene addition mechanism was used to generate cross-linked, polymeric coatings bearing residual thiol moieties on pre-fabricated poly(ethylene glycol) (PEG)-based hydrogel supports. The photopolymerized coatings were 60 μm thick and contained 0.55 nmol unreacted free thiols, corresponding to a concentration of 410 μM, for use as cecropin A (CPA) immobilization handles via thiol-maleimide conjugation, where the CPA-bound maleimide moiety was localized at either the carboxyl terminus or mid-sequence between Ala22 and Gly23. Surface presentation of the thiol handles was controlled by varying the thiolated PEG monomer (PEGSH) used in the photopolymerizable formulation. Bactericidal activity of CPA functionalized hydrogels against E. coli K235 indicated that CPA immobilized at the carboxyl terminus killed 94±6% of the inoculated pathogens when coatings were prepared with high molecular weight PEGSH and 99±1% when prepared with low molecular weight PEGSH. E. coli cell death demonstrated a stronger dependence on peptide concentration than PEG linker length or degree of thiol functionalization, with activity ranging from 34±13% to 99±1% bacterial cells killed as the pre-functionalization thiol concentration in the coatings was increased from 90 μM to 990 μM. Finally, the immobilization site on the surface-bound CPA strongly affected antibacterial activity; when mid-sequence modified CPA was bound to a hydrogel coating bearing 990 μM thiol, only 20±4% of the E. coli population was killed.
Instrument
J-1500
Keywords
Circular dichroism, Secondary structure, Biochemistry, Materials