The biological significance of the carbohydrate-binding activities found for pancreatic enzymes in secretion and digestion

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Novel carbohydrate-binding activities have been discovered for principal pancreatic enzymes. Mammalian a-amylases, trypsins, trypsinogen, chymotrypsin, elastase, lipase, and ribonuclease were found to share the binding activity toward N-linked oligosaccharides of glycoproteins, which is different from the substrate recognition. Trypsins were commonly shown by interaction analyses using surface plasmon resonance (SPR) to bind with glycoproteins possessing N-glycans, but not 0-linked mucin-type glycans, while trypsinogen was found to bind to the gycoproteins possessing 0-glycans as well as N -glycans. The binding constants (KA) between trypsins and glycoproteins were 106-1010 M-1 and Ka of trypsin toward fetuin was decreased from101 M-1 to 104M" by deN-glycosylation of fetuin. Binding with biotin-polymer (BP-) sugar probes revealed that each pancreatic enzyme exhibits specific carbohydrate-binding activity with slight differences among their specificities and the glycoprotein-binding activities were due to the affinity of the enzymes toward the component sugars of Nglycans or 0-glycans. The binding activity was found to be unique to pancreatic enzymes and not observed for a-amylase from human saliva, plants, and fungus and lipase from fungi suggesting a pancreas- specific function of the activity. The binding of glycoproteins or carbohydrates enhanced the enzyme activity noncompetitively or uncompetitively, indicating that the recognition site for N-glycans is different from its catalytic site. Binding studies with the BBM fraction indicated the presence of glycoreceptors for each enzyme that serve as scaffolds to achieve an organized and systematic digestion. The sites and the modes of the carbohydrate-binding of pancreatic enzymes are under investigation by using the software developed for the prediction of the carbohydrate-binding site of proteins and the experimental analyses by x-ray crystallography.