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Thunder-DDA-PASEF boosted by MS2rescore enables high-coverage immunopeptidomics and identifies HLA class-I presented SarsCoV-2 spike protein epitopes

ABSTRACT: Human leukocyte antigen (HLA) class I peptide ligands (HLAIps) are key targets for developing vaccines and immunotherapies against infectious pathogens or cancer cells. Identifying HLAIps is challenging due to their high diversity, low abundance, and patient individuality. Here, we developed a highly sensitive method for identifying HLAIps using liquid chromatography-ion mobility-tandem mass spectrometry (LC-IMS-MS/MS). The optimized method, Thunder-DDA-PASEF, semi-selectively fragments HLAIps based on their IMS and m/z, thus increasing the coverage of immunopeptidomics analyses. Thunder-DDA-PASEF includes singly-charged peptides, which contribute to more than 35% of the HLAIp identifications. Combined with MS2Rescore, Thunder-DDA-PASEF improved ligandome coverage by 150% compared to the Standard-DDA-PASEF method, and enabled in-depth profiling of HLAIps from two human cell lines, JY and Raji, transfected to express the SARS-CoV-2 spike protein. We identified 16 spike protein HLAIps, thirteen of which had been reported to elicit immune responses in human patients.

INSTRUMENT(S): timsTOF Pro 2

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Jeewan Babu Rijal

LAB HEAD: Christine Carapito

PROVIDER: PXD046543 | Pride | 2024-03-19

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	HelaPierceRawFiles.zip	Other
	Human_DCpSP_QC-instru_10022023.fasta	Fasta
	ProlineValidationFiles.zip	Other
	SDRF.tsv	Tabular
	dat files.zip	Other

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Thunder-DDA-PASEF boosted by MS2rescore enables high-coverage immunopeptidomics and identifies HLA class-I presented SarsCoV-2 spike protein epitopes

Project description:Human leukocyte antigen (HLA) class I peptide ligands (HLAIps) are key targets for developing vaccines and immunotherapies against infectious pathogens or cancer cells. Identifying HLAIps is challenging due to their high diversity, low abundance, and patient individuality. Here, we developed a highly sensitive method for identifying HLAIps using liquid chromatography-ion mobility-tandem mass spectrometry (LC-IMS-MS/MS). The optimized method, Thunder-DDA-PASEF, semi-selectively fragments HLAIps based on their IMS and m/z, thus increasing the coverage of immunopeptidomics analyses. Thunder-DDA-PASEF includes singly-charged peptides, which contribute to more than 35% of the HLAIp identifications. Combined with MS2Rescore, Thunder-DDA-PASEF improved ligandome coverage by 150% compared to the Standard-DDA-PASEF method, and enabled in-depth profiling of HLAIps from two human cell lines, JY and Raji, transfected to express the SARS-CoV-2 spike protein. We identified 16 spike protein HLAIps, thirteen of which had been reported to elicit immune responses in human patients.

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Thunder-DDA-PASEF enables high-coverage immunopeptidomics and is boosted by MS2Rescore with novel MS2PIP timsTOF fragmentation prediction model

Project description:Human leukocyte antigen (HLA) class I peptide ligands (HLAIps) are key targets for developing vaccines and immunotherapies against infectious pathogens or cancer cells. Identifying HLAIps is challenging due to their high diversity, low abundance, and patient individuality. Here, we developed a highly sensitive method for identifying HLAIps using liquid chromatography-ion mobility-tandem mass spectrometry (LC-IMS-MS/MS). In addition, we trained a novel timsTOF-specific peak intensity MS2PIP model for tryptic and non-tryptic peptides and implemented it in MS2Rescore (v3) together with the CCS predictor from ionmob. The optimized method, Thunder-DDA-PASEF, semi-selectively fragments singly and multiply charged HLAIps based on their IMS and m/z. Moreover, the method employs the high sensitivity mode and extended IMS resolution with fewer MS/MS frames (300 ms TIMS ramp, 3 MS/MS frames), doubling the coverage of immunopeptidomics analyses, compared to the proteomics-tailored DDA-PASEF (100 ms TIMS ramp, 10 MS/MS frames). Additionally, rescoring boosted the HLAIps identification by 41.7% to 33%. This enabled in-depth profiling of HLAIps from diverse human cell lines and human plasma. Finally, profiling JY and Raji cells transfected to express the SARS-CoV-2 spike protein resulted in 16 spike HLAIps, thirteen of which had been reported to elicit immune responses in human patients.

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