title: Rapid and enhanced proteolytic digestion using electric-field-oriented enzyme reactor.
abstract: We have created a novel enzyme reactor using electric field-mediated orientation and immobilization of proteolytic enzymes (trypsin/chymotrypsin) on biocompatible PVDF membranes in a continuous flow-through chamber. Using less than 5min, this reactor in various enzyme combinations can produce enhanced rapid digestion for standardized prototypic proteins, hydrophilic proteins and hydrophobic transmembrane proteins when compared to in-solution techniques. With improved digestive efficiency, our reactor improved the overall functional analysis of lipid raft proteomes by identifying more closely functionally linked proteins and elucidated a richer set of biological processes and pathways linked to the proteins than traditional in-solution methods.
authors.affiliation: Receptor Pharmacology Unit, Laboratory of Neuroscience, National Institute on Aging, Biomedical Research Center, Baltimore, MD, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhou
authors.forename: Tie
authors.initials: T
authors.lastname: Yi
authors.forename: Sung-Soo
authors.initials: SS
authors.lastname: Park
authors.forename: Wayne
authors.initials: W
authors.lastname: Chadwick
authors.forename: Rong-Fong
authors.initials: RF
authors.lastname: Shen
authors.forename: Wells W
authors.initials: WW
authors.lastname: Wu
authors.forename: Bronwen
authors.initials: B
authors.lastname: Martin
authors.forename: Stuart
authors.initials: S
authors.lastname: Maudsley
mesh_terms: D019149:Bioreactors; D002918:Chymotrypsin / Q000378:metabolism; D004560:Electricity; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006868:Hydrolysis; D021962:Membrane Microdomains / Q000378:metabolism; D008567:Membranes, Artificial; D011145:Polyvinyls; D011506:Proteins / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D014357:Trypsin / Q000378:metabolism; D005465:Fluorocarbon Polymers
chemical_list: D002918:Chymotrypsin; D004800:Enzymes, Immobilized; D008567:Membranes, Artificial; D011145:Polyvinyls; D011506:Proteins; D014357:Trypsin; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers