Abstract
Objectives
Vitamin D-binding protein (VDBP), a serum transport protein for 25-hydroxyvitamin D [25(OH)D], has three common proteoforms which have co-localized amino acid variations and glycosylation. A monoclonal immunoassay was found to differentially detect VDBP proteoforms and methods using liquid chromatography-tandem mass spectrometry (LC-MS/MS) might be able to overcome this limitation. Previously developed multiple reaction monitoring LC-MS/MS methods for total VDBP quantification represent an opportunity to probe the potential effects of proteoforms on proteolysis, instrument response and quantification accuracy.
Methods
VDBP was purified from homozygous human donors and quantified using proteolysis or acid hydrolysis and LC-MS/MS. An interlaboratory comparison was performed using pooled human plasma [Standard Reference Material® 1950 (SRM 1950) Metabolites in Frozen Human Plasma] and analyses with different LC-MS/MS methods in two laboratories.
Results
Several shared peptides from purified proteoforms were found to give reproducible concentrations [≤2.7% coefficient of variation (CV)] and linear instrument responses (R2≥0.9971) when added to human serum. Total VDBP concentrations from proteolysis or amino acid analysis (AAA) of purified proteoforms had ≤1.92% CV. SRM 1950, containing multiple proteoforms, quantified in two laboratories resulted in total VDBP concentrations with 7.05% CV.
Conclusions
VDBP proteoforms were not found to cause bias during quantification by LC-MS/MS, thus demonstrating that a family of proteins can be accurately quantified using shared peptides. A reference value was assigned for total VDBP in SRM 1950, which may be used to standardize methods and improve the accuracy of VDBP quantification in research and clinical samples.
Acknowledgments
The authors would like to thank Dr. Stephen Wise and Dr. Adam Kuszak for their advice and critical review of the manuscript.
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Research funding: This work was partially funded by the UW Nutrition Obesity Research Center (P30DK035816), UW Diabetes Research Center (P30DK017047), and the NIH-ODS through an interagency agreement.
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Author contributions: All authors have accepted responsibility for the entire content of this manuscript and approved its submission.
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Competing interests: Authors state no conflict of interest.
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Informed consent: Informed consent was obtained from all individuals included in this study.
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Ethical approval: Research involving human subjects complied with all relevant national regulations, institutional policies and is in accordance with the tenets of the Helsinki Declaration (as revised in 2018), and has been approved by the authors’ Institutional Review Boards (Committee on Medical Ethics at the University Hospitals Leuven and KU Leuven and the NIST Research Protections Office).
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Disclaimer: Certain commercial instruments, software or materials are identified in this document. Such identification does not imply recommendation or endorsement by NIST, nor does it imply that the products identified are necessarily the best available for the purpose.
References
1. Tsuprykov, O, Chen, X, Hocher, CF, Skoblo, R, Lianghong, Y, Hocher, B. Why should we measure free 25(OH) vitamin D? J Steroid Biochem Mol Biol 2018;180:87–104. https://doi.org/10.1016/j.jsbmb.2017.11.014.Search in Google Scholar PubMed
2. Chun, RF, Peercy, BE, Orwoll, ES, Nielson, CM, Adams, JS, Hewison, M. Vitamin D and DBP: the free hormone hypothesis revisited. J Steroid Biochem Mol Biol 2014;144:132–7. https://doi.org/10.1016/j.jsbmb.2013.09.012.Search in Google Scholar PubMed PubMed Central
3. Bouillon, R, Schuit, F, Antonio, L, Rastinejad, F. Vitamin D binding protein: a historic overview. Front Endocrinol 2020;10:1–21. https://doi.org/10.3389/fendo.2019.00910.Search in Google Scholar PubMed PubMed Central
4. Henderson, CM, Lutsey, PL, Misialek, JR, Laha, TJ, Selvin, E, Eckfeldt, JH, et al.. Measurement by a novel LC-MS/MS methodology reveals similar serum concentrations of vitamin D-binding protein in blacks and whites. Clin Chem 2016;62:179–87. https://doi.org/10.1373/clinchem.2015.244541.Search in Google Scholar PubMed PubMed Central
5. Kilpatrick, LE, Boggs, ASP, Davis, WC, Long, SE, Yen, JH, Phinney, KW. Assessing a method and reference material for quantification of vitamin D binding protein during pregnancy. Clin Mass Spectrom 2020;16:11–7. https://doi.org/10.1016/j.clinms.2020.01.002.Search in Google Scholar PubMed PubMed Central
6. Kilpatrick, LE, Phinney, KW. Quantification of total vitamin-D-binding protein and the glycosylated isoforms by liquid chromatography-isotope dilution mass spectrometry. J Proteome Res 2017;16:4185–95. https://doi.org/10.1021/acs.jproteome.7b00560.Search in Google Scholar PubMed
7. Hoofnagle, AN, Eckfeldt, JH, Lutsey, PL. Vitamin D-binding protein concentrations quantified by mass spectrometry. N Engl J Med 2015;373:1480–2. https://doi.org/10.1056/nejmc1502602.Search in Google Scholar PubMed PubMed Central
8. May, W, Parris, R, Beck, C, Fassett, J, Greenberg, R, Guenther, F, et al.. Definitions of terms and modes used at NIST for value-assignment of reference materials for chemical measurements. Gaithersburg, MD: U.S. Government Printing Office; 2000:1–12 pp.Search in Google Scholar
9. Camara, J, Gonzalez, CA, Choquette, SJ. National institute of standards & technology certificate of analysis standard reference material 1950 metabolites in frozen human plasma. Gaithersburg, MD: NIST Office of Reference Materials; 2019:1–15 pp.Search in Google Scholar
10. Russell, A, Gonzalez, CA, Choquette, SJ. National institute of standards & technology certificate of analysis standard reference material 1949 frozen human prenatal serum. Gaithersburg, MD: NIST Office of Reference Materials; 2021:1–8 pp.Search in Google Scholar
11. Hatse, S, Lambrechts, D, Verstuyf, A, Smeets, A, Brouwers, B, Vandorpe, T, et al.. Vitamin D status at breast cancer diagnosis: correlation with tumor characteristics, disease outcome, and genetic determinants of vitamin D insufficiency. Carcinogenesis 2012;33:1319–26. https://doi.org/10.1093/carcin/bgs187.Search in Google Scholar PubMed
12. Magrane, M, Consortium, U. UniProt knowledgebase: a hub of integrated protein data. J Biol Databases Curation 2011;2011:bar009. https://doi.org/10.1093/database/bar009.Search in Google Scholar PubMed PubMed Central
13. Hoofnagle, AN, Whiteaker, JR, Carr, SA, Kuhn, E, Liu, T, Massoni, SA, et al.. Recommendations for the generation, quantification, storage, and handling of peptides used for mass spectrometry-based assays. Clin Chem 2016;62:48–69. https://doi.org/10.1373/clinchem.2015.250563.Search in Google Scholar PubMed PubMed Central
14. Lowenthal, MS, Yen, J, Bunk, DM, Phinney, KW. Certification of NIST standard reference material 2389a, amino acids in 0.1 mol/L HCl--quantification by ID LC-MS/MS. Anal Bioanal Chem 2010;397:511–9. https://doi.org/10.1007/s00216-010-3616-9.Search in Google Scholar PubMed
15. Taylor, BN, Kuyatt, CE. NIST guidelines for evaluating and expressing the uncertainty of NIST measurement results. Gaithersburg, MD: U.S. Department of Commerce; 1994. Available from: https://www.nist.gov/pml/nist-technical-note-1297.10.6028/NIST.TN.1297Search in Google Scholar
16. Working Group 1 of the Joint Committee for Guides in Metrology. Evaluation of measurement data — guide to the expression of uncertainty in measurement (ISO GUM 1995 with minor corrections). JCGM 100:2008 2008; 1–120. Available from: http://www.bipm.org/utils/common/documents/jcgm/JCGM_100_2008_E.pdf.Search in Google Scholar
17. Ravnsborg, T, Olsen, DT, Thysen, AH, Christiansen, M, Houen, G, Hojrup, P. The glycosylation and characterization of the candidate Gc macrophage activating factor. Biochim Biophys Acta 2010;1804:909–17. https://doi.org/10.1016/j.bbapap.2009.12.022.Search in Google Scholar PubMed
18. Borges, CR, Jarvis, JW, Oran, PE, Nelson, RW. Population studies of vitamin D binding protein microheterogeneity by mass spectrometry lead to characterization of its genotype-dependent O-glycosylation patterns. J Proteome Res 2008;7:4143–53. https://doi.org/10.1021/pr8002936.Search in Google Scholar PubMed
19. Borges, CR, Jarvis, JW, Oran, PE, Rogers, SP, Nelson, RW. Population studies of intact vitamin D binding protein by affinity capture ESI-TOF-MS. J Biomol Tech JBT 2008;19:167–76.Search in Google Scholar
20. Rehder, DS, Nelson, RW, Borges, CR. Glycosylation status of vitamin D binding protein in cancer patients. Protein Sci 2009;18:2036–42. https://doi.org/10.1002/pro.214.Search in Google Scholar PubMed PubMed Central
21. Yu, L, Vizel, A, Huff, MB, Young, M, Remmele, RLJr, He, B. Investigation of N-terminal glutamate cyclization of recombinant monoclonal antibody in formulation development. J Pharmaceut Biomed Anal 2006;42:455–63. https://doi.org/10.1016/j.jpba.2006.05.008.Search in Google Scholar PubMed
22. CLSI. Liquid chromatography-mass spectrometry methods; approved guideline. Wayne, PA: Clinical and Laboratory Standards Institute; 2014.Search in Google Scholar
23. U.S. Department of Health and Human Services Food and Drug Administration. Bioanalytical method validation guidance for industry: food and drug administration; 2018. Available from: https://www.fda.gov/media/70858/download.Search in Google Scholar
24. Shuford, CM, Sederoff, RR, Chiang, VL, Muddiman, DC. Peptide production and decay rates affect the quantitative accuracy of protein cleavage isotope dilution mass spectrometry (PC-IDMS). Mol Cell Proteomics 2012;11:814–23. https://doi.org/10.1074/mcp.o112.017145.Search in Google Scholar PubMed PubMed Central
25. Grangeon, A, Clermont, V, Barama, A, Gaudette, F, Turgeon, J, Michaud, V. Development and validation of an absolute protein assay for the simultaneous quantification of fourteen CYP450s in human microsomes by HPLC-MS/MS-based targeted proteomics. J Pharmaceut Biomed Anal 2019;173:96–107. https://doi.org/10.1016/j.jpba.2019.05.006.Search in Google Scholar PubMed
26. Trenchevska, O, Nelson, RW, Nedelkov, D. Mass spectrometric immunoassays for discovery, screening and quantification of clinically relevant proteoforms. Bioanalysis 2016;8:1623–33. https://doi.org/10.4155/bio-2016-0060.Search in Google Scholar PubMed PubMed Central
27. Zhou, H, Hoek, M, Yi, P, Rohm, RJ, Mahsut, A, Brown, P, et al.. Rapid detection and quantification of apolipoprotein L1 genetic variants and total levels in plasma by ultra-performance liquid chromatography/tandem mass spectrometry. Rapid Commun Mass Spectrom : RCM 2013;27:2639–47. https://doi.org/10.1002/rcm.6734.Search in Google Scholar PubMed
28. Vilà-Rico, M, Colomé-Calls, N, Martín-Castel, L, Gay, M, Azorín, S, Vilaseca, M, et al.. Quantitative analysis of post-translational modifications in human serum transthyretin associated with familial amyloidotic polyneuropathy by targeted LC-MS and intact protein MS. J Proteonomics 2015;127:234–46. https://doi.org/10.1016/j.jprot.2015.04.016.Search in Google Scholar PubMed
29. Little, RR, Rohlfing, CL, Hanson, S, Connolly, S, Higgins, T, Weykamp, CW, et al.. Effects of hemoglobin (Hb) E and HbD traits on measurements of glycated Hb (HbA1c) by 23 methods. Clin Chem 2008;54:1277–82. https://doi.org/10.1373/clinchem.2008.103580.Search in Google Scholar PubMed
30. Shuford, CM, Walters, JJ, Holland, PM, Sreenivasan, U, Askari, N, Ray, K, et al.. Absolute protein quantification by mass spectrometry: not as simple as advertised. Anal Chem 2017;89:7406–15. https://doi.org/10.1021/acs.analchem.7b00858.Search in Google Scholar PubMed
31. van den Broek, I, Romijn, FP, Smit, NP, van der Laarse, A, Drijfhout, JW, van der Burgt, YE, et al.. Quantifying protein measurands by peptide measurements: where do errors arise? J Proteome Res 2015;14:928–42. https://doi.org/10.1021/pr5011179.Search in Google Scholar PubMed
32. Kritmetapak, K, Pongchaiyakul, C. Parathyroid hormone measurement in chronic kidney disease: from basics to clinical implications. Internet J Nephrol 2019;2019:5496710. https://doi.org/10.1155/2019/5496710.Search in Google Scholar PubMed PubMed Central
33. Smit, MA, van Kinschot, CMJ, van der Linden, J, van Noord, C, Kos, S. Clinical guidelines and PTH measurement: does assay generation matter? Endocr Rev 2019;40:1468–80. https://doi.org/10.1210/er.2018-00220.Search in Google Scholar PubMed
Supplementary Material
The online version of this article offers supplementary material (https://doi.org/10.1515/cclm-2022-0642).
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