Hands-On Electrospray Ionization Mass Spectrometry for Undergraduate Biochemistry Students: Peptide Identification by Ladder Sequencing

• Viveka Perera ^[1] ; Baku Acharya ^[1] ; Amanda L. Patrick ^[1] ; Deb Mlsna ^[1]
  1. [1] Mississippi State University

     Mississippi State University

     Estados Unidos

     
• Localización: Journal of chemical education, ISSN 0021-9584, Vol. 97, Nº 5, 2020, págs. 1437-1442
• Idioma: inglés
• Texto completo no disponible (Saber más ...)
• Resumen

  • Mass spectrometers are ever-increasingly powerful, user-friendly, and affordable. Thus, the addition of mass spectrometry experiments into the undergraduate laboratory curriculum is now both feasible and an effective tool to introduce students to relevant instrumentation. Here an experiment demonstrating the use of a high-resolution electrospray ionization mass spectrometer (ESI-HRMS) for the identification of peptide sequences and the differentiation of isomeric peptides was developed and implemented in a biochemistry laboratory course for third-year chemistry major undergraduate students. This lab activity was a first introduction to mass spectrometry for undergraduate biochemistry students. As a prelab assignment, students were tasked to predict the mass-to-charge ratios and the fragmentation patterns for six given peptides using an online fragmentation prediction tool (Protein Prospector’s MS-Product tool). Students then analyzed two unknown peptides using ESI-HRMS. The theoretical and experimental results were compared to reveal the identity of the two unknown peptides. The student success rate of recognizing the unknown peptide sequences was 87.5%. This laboratory experiment provided students with hands-on experience using a research-grade ESI-HRMS instrument to solve a bioanalytical problem—specifically, the identification of the primary structure of a peptide of unknown sequence. Learning outcomes were evaluated for this experiment, which showed student understanding of peptide sequencing using mass spectrometry.