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Glycoprotein changes occur in not only protein abundance but also the occupancy of each glycosylation site by different glycoforms during biological or pathological processes. Recent advances in mass spectrometry instrumentation and techniques have facilitated analysis of intact glycopeptides in complex biological samples by allowing the users to generate spectra of intact glycopeptides with glycans attached to each specific glycosylation site. However, assigning these spectra, leading to identification of the glycopeptides, is challenging. Here, we report an algorithm, named GPQuest, for site-specific identification of intact glycopeptides using higherenergy collisional dissociation (HCD) fragmentation of complex samples. In this algorithm, a spectral library of glycosite-containing peptides in the sample was built by analyzing the isolated glycosite-containing peptides using HCD LC-MS/MS. Spectra of intact glycopeptides were selected by using glycan oxonium ions as signature ions for glycopeptide spectra. These oxoniumion-containing spectra were then compared with the spectral library generated from glycositecontaining peptides, resulting in assignment of each intact glycopeptide MS/MS spectrum to a specific glycosite-containing peptide. The glycan occupying each glycosite was determined by matching the mass difference between the precursor ion of intact glycopeptide and the glycositecontaining peptide to a glycan database. Using GPQuest, we analyzed LC-MS/MS spectra of protein extracts from prostate tumor LNCaP cells. Without enrichment of glycopeptides from global tryptic peptides and at a false discovery rate of 1%, 1008 glycan-containing MS/MS spectra were assigned to 769 unique intact N-linked glycopeptides, representing 344 N-linked glycosites with 57 different N-glycans. Spectral library matching using GPQuest assigns the HCD LC-MS/MS generated spectra of intact glycopeptides in an automated and high-throughput manner. Additionally, spectral library matching gives the user the possibility of identifying novel or * Corresponding Author. hzhang32@jhmi.edu. ASSOCIATED CONTENT Supporting InformationSupporting information tables contain the experimental spectral library composed for LNCaP cells, the glycan database used in this study, and the intact glycopeptides identified in LNCaP. The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem. 5b00024.The authors declare no competing financial interest. HHS Public AccessAuthor manuscript Anal Chem. Author manuscript; available in PMC 2016 January 21. Published in final edited form as:Anal Chem. 2015 ; 87(10): 5181-5188. doi:10.1021/acs.analchem.5b00024. Author Manuscript Author ManuscriptAuthor ManuscriptAuthor Manuscript modified glycans on specific glycosites that might be missing from the predetermined glycan databases. Graphical abstractGlycosylation is one of the most common protein modifications spanning more that 50% of the proteome. Glycosylation mediates many of the cell functions inclu...

Different shapes of ZnO microcrystals have been achieved controllably by a capping-molecule-assisted hydrothermal process. The flowerlike, disklike, and dumbbell-like ZnO microcrystals of hexagonal phase have been obtained respectively using ammonia, citric acid (CA), and poly(vinyl alcohol) (PVA) as the capping molecules. Only a very strong UV emission at ∼380 nm is observed in the photoluminescence (PL) spectra of the three kinds of ZnO microcrystals, indicative of their high crystal quality. The formation mechanisms for the hydrothermally synthesized microcrystals in different morphologies have been phenomenologically presented.

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