Immobilized residue-specific endoproteinases for protein sequencing.
Before proteins can be sequenced, the peptide chain has to be cut into small fragments of less than about 50 amino acids using residue-specific endoproteinases. These enzymes can be immobilized in a highly active form. Using immobilized endoproteinases for protein sequencing results in a series of advantages: (1) the high enzyme activity in the column results in short reaction times; (2) the protein fragments are easily eluted from the column whilst the endoproteinase is completely retained on the column; the protein fragments are clean yielding in low sequencing background; (3) the protein sample to be sequenced is free of exogenous enzymes; (4) endoproteinase self-digestion is prevented by immobilization; therefore, the sample solution does not contain any endoproteinase fragments; (5) enzymes are especially stable when immobilized. Columns with immobilized endoproteinases can be applied repeatedly and stored for many months.
Ronnenberg
J
J
Mobitec GmbH, Göttingen, Germany.
Preitz
B
B
Wöstemeier
G
G
Diekmann
S
S
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004269:DNA, Bacterial / Q000032:analysis; D010450:Endopeptidases / Q000737:chemistry; D004800:Enzymes, Immobilized; D007328:Insulin / Q000032:analysis; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis; D017421:Sequence Analysis
D004269:DNA, Bacterial; D004800:Enzymes, Immobilized; D007328:Insulin; D010455:Peptides; D010450:Endopeptidases