Trimethylation Enhancement using Diazomethane (TrEnDi): Rapid On-Column Quaternization of Peptide Amino Groups via Reaction with Diazomethane Significantly Enhances Sensitivity in Mass Spectrometry An

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• 作者：Karl V. Wasslen ; Le Hoa Tan ; Jeffrey M. Manthorpe ; Jeffrey C. Smith
• 刊名：Analytical Chemistry
• 出版年：2014
• 出版时间：April 1, 2014
• 年：2014
• 卷：86
• 期：7
• 页码：3291-3299
• 全文大小：497K
• 年卷期：v.86,no.7(April 1, 2014)
• ISSN：1520-6882

文摘

Defining cellular processes relies heavily on elucidating the temporal dynamics of proteins. To this end, mass spectrometry (MS) is an extremely valuable tool; different MS-based quantitative proteomics strategies have emerged to map protein dynamics over the course of stimuli. Herein, we disclose our novel MS-based quantitative proteomics strategy with unique analytical characteristics. By passing ethereal diazomethane over peptides on strong cation exchange resin within a microfluidic device, peptides react to contain fixed, permanent positive charges. Modified peptides display improved ionization characteristics and dissociate via tandem mass spectrometry (MS²) to form strong a₂ fragment ion peaks. Process optimization and determination of reactive functional groups enabled a priori prediction of MS² fragmentation patterns for modified peptides. The strategy was tested on digested bovine serum albumin (BSA) and successfully quantified a peptide that was not observable prior to modification. Our method ionizes peptides regardless of proton affinity, thus decreasing ion suppression and permitting predictable multiple reaction monitoring (MRM)-based quantitation with improved sensitivity.