An optimized protocol for nano-LC-MALDI-TOF-MS coupling for the analysis of proteolytic digests of glycoproteins

Biomedical Chromatography : BMC
Günter Lochnit, R Geyer

Abstract

Matrix-assissted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analyses of complete proteolytic digests are often hampered by contaminations and the complexity of the sample. This results in suppression effects and the formation of adducts which are difficult to assign, thus leading to low scores in database searches. In particular, signals of post-translationally modified peptides such as glycopeptides are often of low intensity or completely suppressed. Online liquid chromatography electrospray ionization mass spectrometry (ESI-MS) can, in part, overcome this problem, but the analytes are completely consumed during the run. Coupling of nano-flow HPLC (nano-LC), microfractionation and MALDI-TOF-MS combines separation and high-sensitivity UV detection with the possibility of collecting fractionated peptides and preserving the sample for detailed mass spectrometric analyses. Here we report on an optimized protocol for nano-LC-MALDI-TOF-MS analyses of glycoproteins. This protocol improves spectral quality, resulting in better protein identification scores in database searches. Furthermore, post-translationally modified peptides could be detected with higher sensitivity by changing the experimental co...Continue Reading

References

May 18, 2000·Analytical Chemistry·A ShevchenkoK G Standing
Mar 20, 2003·Journal of Proteome Research·Tomas RejtarBarry L Karger
Sep 5, 2003·Journal of the American Society for Mass Spectrometry·Wanda M BodnarM Arthur Moseley

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Citations

Oct 11, 2005·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Manfred WuhrerCornelis H Hokke
Dec 1, 2006·Biochimica Et Biophysica Acta·Hildegard Geyer, Rudolf Geyer
Aug 18, 2005·Journal of Mass Spectrometry : JMS
Apr 10, 2009·Mass Spectrometry Reviews·Hyejin HwangJing Zhang
Aug 13, 2015·Journal of Chromatography. a·Jozef ŠestákVladislav Kahle
Jun 22, 2016·Journal of Pharmaceutical and Biomedical Analysis·Maria Kristina ParrHouda Montacir
Jun 8, 2010·Molecular and Cellular Biochemistry·Haige YeXiaoshu Zhao

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