A novel organic-inorganic hybrid monolith for trypsin immobilization.
In proteomics, attention has focused on various immobilized enzyme reactors (IMERs) for the realization of high throughput digestion. In this report, a novel organic-inorganic hybrid monolith based IMER was prepared in a 100 μm i.d. capillary with 3-glycidoxypropyltrimethoxysilane (GLYMO) as the monomer and tetraethoxysilane (TEOS) as the crosslinker. Trypsin immobilization was achieved via the reaction between vicinal diol groups, which were obtained from hydrolysis of epoxy groups, and the amino groups of trypsin. Bovine serum albumin was digested thoroughly by this IMER in 47 s. After micro-reverse phase liquid chromatography-tandem mass spectrometry (μRPLC-MS/MS) analysis and database searching, beyond 35% sequence coverage was obtained, and the result was comparable to that of 12 h in solution digestion. The present IMER has potential for high throughput digestion.
Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
ShuaiBin
S
Wu
JunFeng
J
Ma
KaiGuang
K
Yang
JinXiang
J
Liu
Zhen
Z
Liang
LiHua
L
Zhang
YuKui
Y
Zhang
D000818:Animals; D019149:Bioreactors; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods; D015394:Molecular Structure; D012710:Serum Albumin, Bovine / Q000032:analysis; D053719:Tandem Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D014357:Trypsin / Q000737:chemistry* / Q000378:metabolism
D004800:Enzymes, Immobilized; D012710:Serum Albumin, Bovine; D014357:Trypsin