Online coupling of reverse-phase and hydrophilic interaction liquid chromatography for protein and glycoprotein characterization.

Analytical and Bioanalytical Chemistry
Maggie P Y LamIvan K Chu

Abstract

We have developed a novel system for coupling reverse-phase (RP) and hydrophilic interaction liquid chromatography (HILIC) online in a micro-flow scheme. In this approach, the inherent solvent incompatibility between RP and HILIC is overcome through the use of constant-pressure online solvent mixing, which allows our system to perform efficient separations of both hydrophilic and hydrophobic compounds for mass spectrometry-based proteomics applications. When analyzing the tryptic digests of bovine serum albumin, ribonuclease B, and horseradish peroxidase, we observed near-identical coverage of peptides and glycopeptides when using online RP-HILIC--with only a single sample injection event--as we did from two separate RP and HILIC analyses. The coupled system was also capable of concurrently characterizing the peptide and glycan portions of deglycosylated glycoproteins from one injection event, as confirmed, for example, through our detection of 23 novel glycans from turkey ovalbumin. Finally, we validated the applicability of using RP-HILIC for the analysis of highly complex biological samples (mouse chondrocyte lysate, deglycosylated human serum). The enhanced coverage and efficiency of online RP-HILIC makes it a viable techni...Continue Reading

References

Aug 1, 1997·Analytical Chemistry·T Yoshida
Dec 17, 2003·Analytical Chemistry·Yehia MechrefCheni Krishnan
Sep 4, 2004·Journal of Biochemical and Biophysical Methods·Tatsunari Yoshida
Jul 26, 2005·Rapid Communications in Mass Spectrometry : RCM·Ying Qing YuJohn C Gebler
Oct 11, 2005·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·M C García
Sep 15, 2006·Journal of Separation Science·Petrus Hemström, Knut Irgum
Nov 17, 2007·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Koen SandraPat Sandra
Feb 12, 2008·Analytical and Bioanalytical Chemistry·Paul J BoersemaAlbert J R Heck
Feb 13, 2008·Journal of Separation Science·José María Saz, María Luisa Marina
Mar 4, 2008·Journal of Proteome Research·Alessio CeroniStuart M Haslam

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Citations

Mar 28, 2013·Chemical Reviews·William R AlleyMilos V Novotny
Dec 6, 2012·Bioanalysis·Nm CassianoQb Cass
Jun 4, 2014·Mass Spectrometry Reviews·Yeong Hee AhnJong Shin Yoo
Sep 15, 2015·Metabolomics : Official Journal of the Metabolomic Society·Jennifer HaggartyKarl E V Burgess
Dec 18, 2013·Analytical Sciences : the International Journal of the Japan Society for Analytical Chemistry·Shigeo Suzuki
Jun 9, 2021·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Lana Abou ZeidCarole Bresson
Mar 20, 2018·Chemical Reviews·L Renee RuhaakCarlito B Lebrilla

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Methods Mentioned

BETA
MDS
glycosylation
immunodepletion

Software Mentioned

GlcyoWorkbench
GlycoWorkbench
ProteinPilot
Glycomod
Sequence Specific Retention Calculator
ProGroup

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