Development of a parallel microbore hollow fiber enzyme reactor platform for online 18O-labeling: Application to lectin-specific lung cancer N-glycoproteome.
J Chromatogr B Analyt Technol Biomed Life Sci
; 1100-1101: 58-64, 2018 Nov 15.
Article
in En
| MEDLINE
| ID: mdl-30292950
ABSTRACT
We introduce a simple online 18O-labeling protocol for protein samples that uses a parallelizing microbore hollow fiber enzyme reactor (mHFER) as an alternative tool for online proteolytic digestion. Online 18O-labeling is performed by separately attaching two mHFERs in parallel to a 10-port switching valve with a high-pressure syringe pump and two syringes containing 16O- or 18O-water. 16O-/18O-labeled peptides are formed in this manner and simultaneously analyzed online using nanoflow liquid chromatography-tandem mass spectrometry (nLC-MS/MS) without any residual trypsin activity. The usefulness of a parallel mHFER platform (P-mHFER) in 18O-labeling was tested using both cytochrome C and alpha-1-acid-glycoprotein to verify the incorporation level of two 18O atoms into tryptic peptides and to provide a quantitative assessment with varied mixing ratios. Additionally, our 18O-labeling approach was used to study the serum N-glycoproteome from lung cancer patients and controls to evaluate the applicability of lectin-based quantitative N-glycoproteomics. We successfully quantified 76 peptides (from 62 N-glycoproteins). Nineteen of these peptides from lung cancer serum were up-/down-regulated at least 2.5-fold compared to controls. As a result, the P-mHFER-based online 18O-labeling platform presented here can be a simple and reproducible way to allow quantitative proteomic analysis of diverse proteome samples.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Oxygen Isotopes
/
Glycoproteins
/
Bioreactors
/
Proteome
/
Lung Neoplasms
Limits:
Humans
Language:
En
Journal:
J Chromatogr B Analyt Technol Biomed Life Sci
Journal subject:
ENGENHARIA BIOMEDICA
Year:
2018
Document type:
Article
Country of publication:
HOLANDA
/
HOLLAND
/
NETHERLANDS
/
NL
/
PAISES BAJOS
/
THE NETHERLANDS