Identification and Mitigation of Defensins in the Immunopurification of Peptide MHC-I Antigens from Lung Tissue
- 13 October 2021
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of the American Society for Mass Spectrometry
- Vol. 33 (9), 1590-1597
- https://doi.org/10.1021/jasms.1c00024
Abstract
The class I major histocompatibility (MHC-I) complex is a set of diverse cell surface receptors encoded by the human leukocyte antigen gene complex. These receptors present intracellular antigens to cytotoxic T cells providing information on the state and health of cells. Changes in the immunopeptidome during cancer may provide novel targets for therapeutic intervention. To understand how the tumor immunopeptidome is altered, we developed a mass spectrometry (MS) based platform for isolating and identifying MHC-I peptide antigens in lung tumors. In the course of our work, we encountered several large unknown peptide contaminants which had not been previously reported. To understand the source of these major contaminants, we isolated them using offline fractionation and identified them by liquid chromatography–tandem mass spectrometry (LC–MS/MS) as members of the host defense protein family known as the defensins. To mitigate their detrimental effects, we modified our “Original” data-dependent acquisition (DDA) MS method to narrowly target the MHC-I peptides based on their physical properties including charge state and molecular weight (“z state” DDA), evaluated field asymmetric ion mobility spectrometry to attempt gas-phase separation prior to MS analysis, and developed an immunodepletion approach using defensin specific antibodies. This modified approach improves peptide identification and reduces the impact of defensin contamination in lung tissue samples.Keywords
This publication has 32 references indexed in Scilit:
- Simultaneous alignment and clustering of peptide data using a Gibbs sampling approachBioinformatics, 2012
- α‐Defensins in human innate immunityImmunological Reviews, 2011
- Towards a systems understanding of MHC class I and MHC class II antigen presentationNature Reviews Immunology, 2011
- Defensins in innate immunityCell and tissue research, 2010
- Multivalent Binding of Carbohydrates by the Human α-Defensin, HD5The Journal of Immunology, 2009
- CTdatabase: a knowledge-base of high-throughput and curated data on cancer-testis antigensNucleic Acids Research, 2008
- The MHC class I antigen presentation pathway: strategies for viral immune evasionImmunology, 2003
- Defensins: antimicrobial peptides of innate immunityNature Reviews Immunology, 2003
- Longer peptide can be accommodated in the MHC class I binding site by a protrusion mechanismEuropean Journal of Immunology, 2000
- Isolation and analysis of naturally processed viral peptides as recognized by cytotoxic T cellsNature, 1990