Towards absolute quantification of protein genetic variants in Pisum sativum extracts.

Autor: Vreeke GJC; Laboratory of Food Chemistry, Wageningen University and Research, P.O. Box 17, 6700 AA, Wageningen, the Netherlands., Meijers MGJ; Laboratory of Food Chemistry, Wageningen University and Research, P.O. Box 17, 6700 AA, Wageningen, the Netherlands; TiFN, P.O. Box 557, 6700 AN, Wageningen, the Netherlands., Vincken JP; Laboratory of Food Chemistry, Wageningen University and Research, P.O. Box 17, 6700 AA, Wageningen, the Netherlands., Wierenga PA; Laboratory of Food Chemistry, Wageningen University and Research, P.O. Box 17, 6700 AA, Wageningen, the Netherlands. Electronic address: peter.wierenga@wur.nl.
Jazyk: angličtina
Zdroj: Analytical biochemistry [Anal Biochem] 2023 Mar 15; Vol. 665, pp. 115048. Date of Electronic Publication: 2023 Jan 16.
DOI: 10.1016/j.ab.2023.115048
Abstrakt: In recent years, several studies have used proteomics approaches to characterize genetic variant profiles of agricultural raw materials. In such studies, the challenge is the quantification of the individual protein variants. In this study a novel UPLC-PDA-MS method with absolute and label-free UV-based peptide quantification was applied to quantify the genetic variants of legumin, vicilin and albumins in pea extracts. The aim was to investigate the applicability of this method and to identify challenges in determining protein concentration from the measured peptide concentrations. Analysis of the protein mass balance showed significant losses of proteins in extraction (37%) and of peptides in further sample preparation (69%). The challenge in calculating the extractable individual protein concentrations was how to deal with these insoluble peptides. The quantification approach using average amino acid concentrations in each position of the sequence showed most reproducible results and allowed comparison of the genetic protein composition of 8 different cultivars. The extractable protein composition (μM/μM) was remarkably similar for all cultivar extracts and consisted of legumins A1 (12.8 ± 1.2%), A2 (1.1 ± 0.4%), B (9.9 ± 1.6%), J (7.5 ± 1.0%) and K (10.3 ± 2.1%), vicilin (15.2 ± 1.7%), provicilin (15.7 ± 2.5%), convicilin (9.8 ± 0.8%), albumin A1 (7.4 ± 2.0%), albumin 2 (10.0 ± 1.5%) and protease inhibitor (0.4 ± 0.4%).
Competing Interests: Declaration of competing interest The project is partially organized by and executed under the auspices of TiFN, a public - private partnership on precompetitive research in food and nutrition. The authors have declared that no competing interests exist in the writing of this publication. Funding for this research was partially obtained from Bel S.A., Nutricia Research B.V., Pepsico Inc., Unilever Nederland holdings B.V., the Netherlands Organisation for Scientific Research and the Top-sector Agri&Food.
(Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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