Pyridinium-Based Strategy for a Bioorthogonal Conjugation-Assisted Purification Method for Profiling Cell Surface...

Area of Science:

• Biochemistry
• Cell Biology
• Proteomics

Background:

• Cell surface proteins (CSPs) are crucial therapeutic targets, but selective enrichment remains challenging.
• Existing labeling methods often lack specificity, leading to intracellular protein contamination.

Purpose of the Study:

• To develop a novel cross-linking probe for highly selective cell surface protein (CSP) labeling and enrichment.
• To demonstrate the probe's efficiency and specificity in live cell imaging and proteomic analysis.

Main Methods:

• Development and application of a novel sulfo-pyridinium ester (SPE) cross-linking probe.
• Two-step imaging and enrichment strategy for cell surface proteins.
• Proteomic analysis of labeled proteins in MCF-7 cells, including insulin-cultured conditions.

Main Results:

• The SPE probe exhibited superior protein labeling efficiency and Lys specificity compared to NHS esters.
• Selective labeling of cell membranes with minimal intracellular protein labeling was achieved.
• 425 unique CSPs were identified from 1162 labeled proteins in MCF-7 cells.
• Identification of cell surface targets related to functional biomarkers and insulin-associated pathogenesis.

Conclusions:

• The SPE probe offers a promising tool for selective cell surface protein labeling.
• This method facilitates the monitoring of transient cell surface events and aids in identifying therapeutic targets.