A flow-through trypsin bioreactor for peptide fingerprinting based on nanoporous alumina.
(English)Manuscript (preprint) (Other academic)
Trypsin was immobilized on nanoporous anodized alumina membranes to create an enzyme reactor suitable for peptide mass fingerprinting. The membranes were derivatized with 3-aminopropyltriethoxysilane and the amino groups were activated with carbonyl diimidazole to allow coupling of trypsin via e-amino groups. The function was assessed for the artificial substrate BAPA, bovine ribonuclease A and bovine serum albumin. The ribonuclease A was correctly identified from the peptide pattern by a Mascot database search. The activity in a 10-membrane stack was in the range required for on-line ESI-MS peptide mass fingerprinting. The reactor was found to retain 76% of the initial activity after 14 days of storage and repeated use at room temperature.
nanoporous aluminum oxide, immobilization, trypsin, peptide mass fingerprinting, enzyme stability
Biochemistry and Molecular Biology
IdentifiersURN: urn:nbn:se:uu:diva-210118OAI: oai:DiVA.org:uu-210118DiVA: diva2:661109