In Situ Simultaneous Protein−Polysaccharide Bioconjugation and Hydrogelation Using Horseradish Peroxidase

We propose the peroxidase-catalyzed simultaneous conjugation and hydrogelation of polysaccharide and protein derivatives, each possessing phenolic hydroxyl (Ph) moieties, as a novel route for obtaining protein-polysaccharide conjugate hydrogels. We used alginate, gelatin, and albumin derivatives bearing Ph moieties (Alg-Ph, Gela-Ph, and Alb-Ph) to demonstrate the feasibility. The gelation time of conjugate gels decreased with decreasing H(2)O(2) concentration and with increasing horseradish peroxidase concentration. Gelation time was controllable from a few seconds to 6 min. The repulsion force detected at 40% compression of a conjugate gel obtained from a mixture of Alg-Ph and Gela-Ph at 1.0% (w/v), respectively, was more than 2.8 times larger than that detected for gels produced from 3.0% (w/v) Gela-Ph or 2.0% (w/v) Alg-Ph alone. Cell adhesiveness of gels was tunable by changing the type of protein derivative. A gel from Gela-Ph and Alg-Ph showed higher cell adhesiveness than Alg-Ph gel, but a gel produced from Alb-Ph and Alg-Ph showed a lower cell adhesiveness than Alg-Ph gel. The conjugate gel was degradable by degrading alginate molecules using the nonproteolytic enzyme alginate lyase. The tunable gelation, mechanical properties, and cell adhesiveness of polysaccharide-protein conjugate hydrogels obtained through peroxidase-catalyzed gelation indicates great potential for a wide range of applications, such as scaffolds for tissue engineering and carriers for drug delivery system.