Dataset Information

Complementary Proteome and Glycoproteome Access Revealed Through Comparative Analysis of Reversed Phase and Porous Graphitic Carbon Chromatography

ABSTRACT: Aided by improvements in instrumental and analytical techniques, rigorous connections have been drawn between glycoprotein expression, modification heterogeneity, and a variety of human illnesses. These illnesses, such as various forms of cancer, are often associated with poor prognoses, prompting the need for more comprehensive characterization of the glycoproteome. Instrumental advancements, optimal and nascent dissociation techniques, and computational strategies present the largest area of glycoprotein profiling innovation, often neglecting potential benefits provided by alternative chromatography techniques. Porous graphitic carbon (PGC) represents a separation regime that has gained significant interest in glycomics research due to its mobile phase flexibility, increased retention of polar analytes and improved structural elucidation at higher temperatures. PGC has yet to be systematically compared against or in tandem with standard reversed phase liquid chromatography (RPLC) in high-throughput bottom-up glycoproteomics experiments, leaving the potential benefits unexplored. Performing comparative analysis of single and biphasic separation regimes at a range of column temperatures illustrates the advantages for each method. PGC separation is shown to selectively retain shorter, more hydrophilic glycopeptide species, imparting higher average charge, and exhibiting greater microheterogeneity coverage for identified glycosites. Additionally, we demonstrate that liquid-phase separation of glycopeptide isomers may be achieved when in both single and biphasic PGC separation, providing a means towards facile, multiplex glycopeptide characterization. Beyond this, we demonstrate how utilization of multiple separation regimes and column temperatures can aid in profiling the glycoproteome in tumorigenic and aggressive prostate cancer cells.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human) Bos Taurus (bovine)

TISSUE(S): Cell Culture

SUBMITTER: Daniel Delafield

LAB HEAD: Lingjun Li

PROVIDER: PXD024195 | Pride | 2022-02-13

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	20201115_GD_EnrGly_1115_BP_30C_run1.mzXML	Mzxml
	20201115_GD_EnrGly_1115_BP_30C_run1.raw	Raw
	20201115_GD_EnrGly_1115_BP_45C_run1.mzXML	Mzxml
	20201115_GD_EnrGly_1115_BP_45C_run1.raw	Raw
	20201115_GD_EnrGly_1115_BP_60C_run1.mzXML	Mzxml

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Publications

Complementary proteome and glycoproteome access revealed through comparative analysis of reversed phase and porous graphitic carbon chromatography.

Delafield Daniel G DG Miles Hannah N HN Liu Yuan Y Ricke William A WA Li Lingjun L

Analytical and bioanalytical chemistry 20220209 18

Continual developments in instrumental and analytical techniques have aided in establishing rigorous connections between protein glycosylation and human illness. These illnesses, such as various forms of cancer, are often associated with poor prognoses, prompting the need for more comprehensive characterization of the glycoproteome. While innovative instrumental and computational strategies have largely benefited glycoproteomic analyses, less attention is given to benefits gained through alterna ...[more]

PMID: 35137243

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