In vitro protein-polysaccharide conjugation: tyrosinase-catalyzed conjugation of gelatin and chitosan.
The enzyme tyrosinase was used for the in vitro conjugation of the protein gelatin to the polysaccharide chitosan. Tyrosinases are oxidative enzymes that convert accessible tyrosine residues of proteins into reactive o-quinone moieties. Spectrophotometric and dissolved oxygen studies indicate that tyrosinase can oxidize gelatin and we estimate that 1 in 5 gelatin chains undergo reaction. Oxidized tyrosyl residues (i.e., quinone residues) can undergo nonenzymatic reactions with available nucleophiles such as the nucleophilic amino groups of chitosan. Ultraviolet/visible, (1)H-NMR, and ir provided chemical evidence for the conjugation of oxidized gelatin with chitosan. Physical evidence for conjugation was provided by dynamic viscometry, which indicated that tyrosinase catalyzes the sol-to-gel conversion of gelatin/chitosan mixtures. The gels formed from tyrosinase-catalyzed reactions were observed to differ from gels formed by cooling gelatin. In contrast to gelatin gels, tyrosinase-generated gels had different thermal behavior and were broken by the chitosan-hydrolyzing enzyme chitosanase. These results demonstrate that tyrosinase can be exploited for the in vitro formation of protein-polysaccharide conjugates that offer interesting mechanical properties.
Center for Agricultural Biotechnology, University of Maryland Biotechnology Institute, 5115 Plant Sciences Building, College Park, MD 20742-4450, USA.
Tianhong
T
Chen
Heather D
HD
Embree
Li-Qun
LQ
Wu
Gregory F
GF
Payne
D000363:Agaricales; D000818:Animals; D003386:Brachyura; D002686:Chitin / Q000031:analogs & derivatives / Q000737:chemistry*; D048271:Chitosan; D005780:Gelatin / Q000737:chemistry*; D006023:Glycoproteins / Q000737:chemistry*; D006026:Glycoside Hydrolases / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D014442:Monophenol Monooxygenase / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D013053:Spectrophotometry; D017550:Spectroscopy, Fourier Transform Infrared; D013552:Swine; D013696:Temperature
D006023:Glycoproteins; D002686:Chitin; D005780:Gelatin; D048271:Chitosan; D014442:Monophenol Monooxygenase; D006026:Glycoside Hydrolases; C019868:chitosanase