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| Analysis of allergen protein in Macrobrachium by ultra-high performanceliquid chromatography coupled with quadrupole orbitrap mass spectrometry |
| PAN Xiaodong, HUANG Baifen, CAI Zengxuan, WANG Mengli, HAN Jianlong, CHEN Qing
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| Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, Zhejiang 310051, China |
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Abstract Objective To establish the ultra-high performance liquid chromatography coupled with quadrupole orbitrap mass spectrometry ( UPLC-Q-Orbitrap-MS ) for the analysis of allergen protein in Macrobrachium. Methods Based on the strategy of bottom-up protein analysis, the proteins in Macrobrachium samples were extracted by Tris-HCl, digested by trypsin at 40 ℃ for 6 hours, separated by chromatography, and analyzed by Q-Orbitrap-MS spectrometry ( Full MS/dd-MS2, TopN=10 ) . Allergen proteins were identified with UniProt protein database and Proteome Discoverer software. Results Four kinds of allergen proteins were obtained, which were tropomyosin, arginine kinase, sarcoplasmic calcium binding protein and hemocyanin. The coverage rates of peptides in proteins were 53%, 36%, 12% and 12%, respectively. Post translation modifications were methylation of aspartic acid (D), deacylylation of aspartic acid ( N ) , glutamyl ammonia ( Q ) and oxidation of methionine ( M ) . Conclusions The UPLC-Q-Orbitrap-MS can identified abundant peptide and fragment information with high sensitivity and resolution, which provides a technical support for the analysis of shrimp allergens.
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Received: 12 May 2020
Revised: 15 June 2020
Published: 30 September 2020
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