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Terpolymeric platform with enhanced hydrophilicity via cysteic acid for serum intact glycopeptide analysis

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Abstract

A new polymeric (methyl methacrylate/ethylene glycol dimethacrylate/1,2-epoxy-5-hexene) base/matrix has been fabricated and decorated with zwitterionic hydrophilic cysteic acid (Cya) for the enrichment of intact N-glycopeptides from standards and biological samples. Terpolymer-Cya provides good enrichment efficiency, improved hydrophilicity, and selectivity by virtue of better surface area (2.09 × 102 m2/g) provided by terpolymer and the zwitterionic property offered by cysteic acid. Cysteic acid-functionalized polymeric hydrophilic interaction liquid chromatography (HILIC) sorbent enriches 35 and 24 N-linked glycopeptides via SPE (solid phase extraction) mode from tryptic digests of model glycoproteins, i.e., immunoglobulin G (IgG) and horseradish peroxidase (HRP), respectively. Zwitterionic chemistry of cysteine helps in achieving higher selectivity with BSA digest (1:200), and lower detection limit down to 100 attomoles with a complete glycosylation profile of each standard digest. The recovery of 81% and good reproducibility define the application of terpolymer-Cya for complex samples like a serum. Analysis of human serum provides a profile of 807 intact N-linked glycopeptides via nano-liquid chromatography-tandem mass spectrometry (nLC-MS/MS). To the best of our knowledge, this is the highest number of glycopeptides enriched by any HILIC sorbent. Selected glycoproteins are evaluated in link to various cancers including the breast, lung, uterine, and melanoma using single-nucleotide variances (BioMuta). This study represents the complete idea of using an in-house developed strategy as a successful tool to help analyze, relate, and answer glycoprotein-based clinical issues regarding cancers.

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References

  1. Gilgunn S, Conroy PJ, Saldova R, Rudd PM, O’kennedy RJ (2013) Aberrant PSA glycosylation—a sweet predictor of prostate cancer. Nat Rev Urol 10(2):99–107. https://doi.org/10.1038/nrurol.2012.258

    Article  CAS  PubMed  Google Scholar 

  2. Ohtsubo K, Chen MZ, Olefsky JM, Marth JD (2011) Pathway to diabetes through attenuation of pancreatic beta cell glycosylation and glucose transport. Nat Med 17(9):1067–1075. https://doi.org/10.1038/nm.2414

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Wang AC, Jensen EH, Rexach JE, Vinters HV, Hsieh-Wilson LC (2016) Loss of O-GlcNAc glycosylation in forebrain excitatory neurons induces neurodegeneration. Proc Nat Acad Sci 113(52):15120–15125. https://doi.org/10.1073/pnas.1606899113

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Hafkenscheid L, Bondt A, Scherer HU, Huizinga TW, Wuhrer M, Toes RE, Rombouts Y (2017) Structural analysis of variable domain glycosylation of anti-citrullinated protein antibodies in rheumatoid arthritis reveals the presence of highly sialylated glycans. Mol Cell Proteom 16(2):278–287. https://doi.org/10.1074/mcp.M116.062919

    Article  CAS  Google Scholar 

  5. Ghesquière B, Van Damme J, Martens L, Vandekerckhove J, Gevaert K (2006) Proteome-wide characterization of N-glycosylation events by diagonal chromatography. J Proteome Res 5(9):2438–2447. https://doi.org/10.1021/pr060186m

    Article  CAS  PubMed  Google Scholar 

  6. Ruhaak LR, Xu G, Li Q, Goonatilleke E, Lebrilla CB (2018) Mass spectrometry approaches to glycomic and glycoproteomic analyses. Chem Rev 118(17):7886–7930. https://doi.org/10.1021/acs.chemrev.7b00732

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Xiao H, Suttapitugsakul S, Sun F, Wu R (2018) Mass spectrometry-based chemical and enzymatic methods for global analysis of protein glycosylation. Acc Chem Res 51(8):1796–1806. https://doi.org/10.1021/acs.accounts.8b00200

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Liu Q, Deng CH, Sun N (2018) Hydrophilic tripeptide-functionalized magnetic metal–organic frameworks for the highly efficient enrichment of N-linked glycopeptides. Nanoscale 10(25):12149–12155. https://doi.org/10.1039/C8NR03174F

    Article  CAS  PubMed  Google Scholar 

  9. Kailasa SK, D’souza S, Wu HF (2014) Analytical applications of nanoparticles in MALDI-MS for bioanalysis. Bioanalysis 7(17):2265–2276. https://doi.org/10.4155/bio.15.149

    Article  CAS  Google Scholar 

  10. Kailasa S, Wu HF (2014) Recent developments in nanoparticle-based MALDI mass spectrometric analysis of phosphoproteomes. Microchim Acta 181:853–864. https://doi.org/10.1007/s00604-014-1191-z

    Article  CAS  Google Scholar 

  11. Kailasa S, Koduru JK, Baek SH, Wu HF, Hussain CM, Park TJ (2020) Review on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the rapid screening of microbial species: a promising bioanalytical tool. Microchem J 159:105387. https://doi.org/10.1016/j.microc.2020.105387

    Article  CAS  Google Scholar 

  12. Liu L, Yu M, Zhang Y, Wang C, Lu H (2014) Hydrazide functionalized core–shell magnetic nanocomposites for highly specific enrichment of N-glycopeptides. ACS Appl Mater Interfaces 6(10):7823–7832. https://doi.org/10.1021/am501110e

    Article  CAS  PubMed  Google Scholar 

  13. Boersema PJ, Mohammed S, Heck AJ (2008) Hydrophilic interaction liquid chromatography (HILIC) in proteomics. Anal Bioanal Chem 391(1):151–159. https://doi.org/10.1007/s00216-008-1865-7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Tang J, Liu Y, Qi D, Yao G, Deng C, Zhang X (2009) On-plate-selective enrichment of glycopeptides using boronic acid-modified gold nanoparticles for direct MALDI-QIT-TOF MS analysis. Proteomics 9(22):5046–5055. https://doi.org/10.1002/pmic.200900033

    Article  CAS  PubMed  Google Scholar 

  15. Madera M, Mechref Y, Novotny MV (2005) Combining lectin microcolumns with high-resolution separation techniques for enrichment of glycoproteins and glycopeptides. Anal Chem 77(13):4081–4090. https://doi.org/10.1021/ac050222l

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Dong L, Feng S, Li S, Song P, Wang J (2015) Preparation of concanavalin A-chelating magnetic nanoparticles for selective enrichment of glycoproteins. Anal Chem 87(13):6849–6853. https://doi.org/10.1021/acs.analchem.5b01184

    Article  CAS  PubMed  Google Scholar 

  17. Liang Y, Wu C, Zhao Q, Wu Q, Jiang B, Weng Y, Liang Z, Zhang L, Zhang Y (2015) Gold nanoparticles immobilized hydrophilic monoliths with variable functional modification for highly selective enrichment and on-line deglycosylation of glycopeptides. Anal Chim Acta 900:83–89. https://doi.org/10.1016/j.aca.2015.10.024

    Article  CAS  PubMed  Google Scholar 

  18. Qu Y, Sun L, Zhang Z, Dovichi NJ (2018) Site-specific glycan heterogeneity characterization by hydrophilic interaction liquid chromatography solid-phase extraction, reversed-phase liquid chromatography fractionation, and capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry. Anal Chem 90(2):1223–1233. https://doi.org/10.1021/acs.analchem.7b03912

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Saleem S, Sajid MS, Hussain D, Jabeen F, Najam-ul-Haq M, Saeed A (2020) Boronic acid functionalized MOFs as HILIC material for N-linked glycopeptide enrichment. Anal Bioanal Chem 412(7):1509–1520. https://doi.org/10.1007/s00216-020-02427-9

    Article  CAS  PubMed  Google Scholar 

  20. Zhang YW, Li Z, Zhao Q, Zhou YL, Liu HW, Zhang XX (2014) A facilely synthesized amino-functionalized metal–organic framework for highly specific and efficient enrichment of glycopeptides. Chem Commun 50(78):11504–11516. https://doi.org/10.1039/C4CC05179C

    Article  CAS  Google Scholar 

  21. Qing G, Yan J, He X, Li X, Liang X (2020) Recent advances in hydrophilic interaction liquid interaction chromatography materials for glycopeptide enrichment and glycan separation. TrAC Trends Anal Chem 124:115570. https://doi.org/10.1016/j.trac.2019.06.020

    Article  CAS  Google Scholar 

  22. Liu L, Zhang Y, Zhang L, Yan G, Yao J, Yang P, Lu H (2012) Highly specific revelation of rat serum glycopeptidome by boronic acid-functionalized mesoporous silica. Anal Chim Acta 753:64–72. https://doi.org/10.1016/j.aca.2012.10.002

    Article  CAS  PubMed  Google Scholar 

  23. Yu L, Li X, Guo Z, Zhang X, Liang X (2009) Hydrophilic interaction chromatography-based enrichment of glycopeptides by using click maltose: a matrix with high selectivity and glycosylation heterogeneity coverage. Chem Eur J 15(46):12618–12626. https://doi.org/10.1002/chem.200902370

    Article  CAS  PubMed  Google Scholar 

  24. Xiong Z, Qin H, Wan H, Huang G, Zhang Z, Dong J, Zhang L, Zhang W, Zou H (2013) Layer-by-layer assembly of multilayer polysaccharide coated magnetic nanoparticles for the selective enrichment of glycopeptides. Chem Commun 49(81):9284–9296. https://doi.org/10.1039/C3CC45008B

    Article  CAS  Google Scholar 

  25. Bai H, Zhang B, Cheng X, Liu J, Wang X, Qin W, Zhang M (2022) Synthesis of zwitterionic polymer modified graphene oxide for hydrophilic enrichment of N-glycopeptides from urine of healthy subjects and patients with lung adenocarcinoma. Talanta 237:122938. https://doi.org/10.1016/j.talanta.2021.122938

    Article  CAS  PubMed  Google Scholar 

  26. Ma W, Xu L, Li X, Shen S, Wu M, Bai Y, Liu H (2017) Cysteine-functionalized metal–organic framework: facile synthesis and high efficient enrichment of n-linked glycopeptides in cell lysate. ACS Appl Mater Interfaces 9:19562–19568. https://doi.org/10.1021/acsami.7b02853

    Article  CAS  PubMed  Google Scholar 

  27. He Y, Zheng Q, Huang H, Ji Y, Lin Z (2022) Synergistic synthesis of hydrophilic hollow zirconium organic frameworks for simultaneous recognition and capture of phosphorylated and glycosylated peptides. Anal Chim Acta 1198:339552. https://doi.org/10.1016/j.aca.2022.339552

    Article  CAS  PubMed  Google Scholar 

  28. Sajid MS, Jovcevski B, Pukala TL, Jabeen F, Najam-ul-Haq M (2019) Fabrication of piperazine functionalized polymeric monolithic tip for rapid enrichment of glycopeptides/glycans. Anal Chem 92(1):683–689. https://doi.org/10.1021/acs.analchem.9b02068

    Article  CAS  PubMed  Google Scholar 

  29. Shaipulizan NS, Md Jamil SN, Kamaruzaman S, Subri NN, Adeyi AA, Abdullah AH, Abdullah LC (2020) Preparation of ethylene glycol dimethacrylate (EGDMA)-based terpolymer as potential sorbents for pharmaceuticals adsorption. Polymers 12(2):423. https://doi.org/10.3390/polym12020423

    Article  CAS  PubMed Central  Google Scholar 

  30. Demirci S, Khiev D, Can M, Sahiner M, Biswal MR, Ayyala RS, Sahiner N (2021) Chemically cross-linked poly (β-cyclodextrin) particles as promising drug delivery materials. ACS Appl Poly Mater 3:6238–6251. https://doi.org/10.1021/acsapm.1c01058

    Article  CAS  Google Scholar 

  31. Podlesnyuk VV, Hradil J, Marutovskii RM, Klimenko NA, Fridman LE (1997) Sorption of organic compounds from aqueous solutions by glycidyl methacrylate-styrene-ethylene dimethacrylate terpolymers. React Funct Polym 33(2–3):275–288. https://doi.org/10.1016/S1381-5148(97)00065-5

    Article  CAS  Google Scholar 

  32. Miletic N, Vukovic Z, Nastasovic A, Loos K (2009) Macroporous poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) resins—versatile immobilization supports for biocatalysts. J Mol Catal B 56:196–201. https://doi.org/10.1016/j.molcatb.2008.04.012

    Article  CAS  Google Scholar 

  33. Qu JB, Lin YY, Liu Y, Zhu BQ, Sun YJ, Peng WS, Li J (2021) Synthesis of bimodal open-porous polystyrene monoliths with glycopolymer surfaces for high-speed protein chromatography. ACS Appl Polym Mater 3:2657–2666. https://doi.org/10.1021/acsapm.1c00232

    Article  CAS  Google Scholar 

  34. Mompo RO, Vergara BM, Lerma GJ, Simó EF, Herrero M (2021) Boronate affinity sorbents based on thiol-functionalized polysiloxane-polymethacrylate composite materials in syringe format for selective extraction of glycopeptides. Microchem J 164:106018. https://doi.org/10.1016/j.microc.2021.106018

    Article  CAS  Google Scholar 

  35. Furukawa H, Ko N, Go YB, Aratani N, Choi SB, Choi E, Yazaydin AÖ, Snurr RQ, O’Keeffe M, Kim J, Yaghi OM (2010) Ultrahigh porosity in metal-organic frameworks. Science 329(5990):424–428. https://doi.org/10.1126/science.1192160

    Article  CAS  PubMed  Google Scholar 

  36. Diggle B, Jiang Z, Li RW, Connal LA (2021) Self-healing polymer network with high strength, tunable properties, and biocompatibility. Chem Mater 33:3712–3720. https://doi.org/10.1021/acs.chemmater.1c00707

    Article  CAS  Google Scholar 

  37. Ma YF, Yuan F, Zhang XH, Zhou YL, Zhang XX (2017) Highly efficient enrichment of N-linked glycopeptides using a hydrophilic covalent-organic framework. Analyst 142(17):3212–3218. https://doi.org/10.1039/C7AN01027C

    Article  CAS  PubMed  Google Scholar 

  38. Zhang W, Jiang L, Fu L, Jia Q (2019) Selective enrichment of glycopeptides based on copper tetra (N-carbonylacrylic) aminephthalocyanine and iminodiacetic acid functionalized polymer monolith. J Sep Sci 42(5):1037–1044. https://doi.org/10.1002/jssc.201801030

    Article  CAS  PubMed  Google Scholar 

  39. Bi C, Liang Y, Shen L, Tian S, Zhang K, Li Y, He X, Chen L, Zhang Y (2018) Maltose-functionalized hydrophilic magnetic nanoparticles with polymer brushes for highly selective enrichment of N-linked glycopeptides. ACS Omega 3(2):1572–1580. https://doi.org/10.1021/acsomega.7b01788

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Ying LL, Wang DY, Yang HP, Deng XY, Peng C, Zheng C, Xu B, Dong LY, Wang X, Xu L, Zhang YW (2018) Synthesis of boronate-decorated polyethyleneimine-grafted porous layer open tubular capillaries for enrichment of polyphenols in fruit juices. J Chromatogr A 1544:23–32. https://doi.org/10.1016/j.chroma.2018.02.044

    Article  CAS  PubMed  Google Scholar 

  41. Wang YF, Jin HX, Wang YG, Yang LY, OuYang XK, Wu WJ (2016) Synthesis and characterization of magnetic molecularly imprinted polymer for the enrichment of ofloxacin enantiomers in fish samples. Molecules 21(7):915. https://doi.org/10.3390/molecules21070915

    Article  CAS  PubMed Central  Google Scholar 

  42. Zou X, Zhong L, Liu D, Yang B, Lou Y, Peng J, Rainer M, Feuerstein I, Muhammad NUH, Huck CW, Bonn GK (2011) Novel multifunctional chitosan-GMA-IDA-Cu (II) nanospheres for high dynamic range characterization of the human plasma proteome. Anal Bioanal Chem 400(3):747–756. https://doi.org/10.1007/s00216-011-4812-y

    Article  CAS  PubMed  Google Scholar 

  43. Zhao Q, Fang F, Wu C, Wu Q, Liang Y, Liang Z, Zhang L, Zhang Y (2016) imFASP: An integrated approach combining in-situ filter-aided sample pretreatment with microwave-assisted protein digestion for fast and efficient proteome sample preparation. Anal Chim Acta 912:58–64. https://doi.org/10.1016/j.aca.2016.01.049

    Article  CAS  PubMed  Google Scholar 

  44. Peng J, Hu Y, Zhang H, Wan L, Wang L, Liang Z, Zhang L, Wu RA (2019) High anti-interfering profiling of endogenous glycopeptides for human plasma by the dual-hydrophilic metal–organic framework. Anal Chem 91(7):4852–4859. https://doi.org/10.1021/acs.analchem.9b00542

    Article  CAS  PubMed  Google Scholar 

  45. Cao Z, Jiang S (2012) Super-hydrophilic zwitterionic poly (carboxybetaine) and amphiphilic non-ionic poly (ethylene glycol) for stealth nanoparticles. Nano Today 7(5):404–413. https://doi.org/10.1016/j.nantod.2012.08.001

    Article  CAS  Google Scholar 

  46. Sun N, Wu H, Chen H, Shen X, Deng C (2019) Advances in hydrophilic nanomaterials for glycoproteomics. Chem Commun 55(70):10359–10375. https://doi.org/10.1039/C9CC04124A

    Article  CAS  Google Scholar 

  47. Roy R, Ang E, Komatsu E, Domalaon R, Bosseboeuf A, Harb J, Hermouet S, Krokhin O, Schweizer F, Perreault H (2018) Absolute quantitation of glycoforms of two human IgG subclasses using synthetic fc peptides and glycopeptides. J Am Soc Mass Spectrom 29(6):1086–1098. https://doi.org/10.1007/s13361-018-1900-7

    Article  CAS  PubMed  Google Scholar 

  48. Krokhin O, Ens W, Standing KG, Wilkins J, Perreault H (2004) Site-specific N-glycosylation analysis: matrix-assisted laser desorption/ionization quadrupole-quadrupole time-of-flight tandem mass spectral signatures for recognition and identification of glycopeptides. Rapid Commun Mass Spectrom 18(18):2020–2030. https://doi.org/10.1002/rcm.1585

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Wang H, Jiao F, Gao F, Huang J, Zhao Y, Shen Y, Zhang Y, Qian X (2017) Facile synthesis of magnetic covalent organic frameworks for the hydrophilic enrichment of N-glycopeptides. J Mater Chem B 5(22):4052–4059. https://doi.org/10.1039/C7TB00700K

    Article  CAS  PubMed  Google Scholar 

  50. Ji Y, Xiong Z, Huang G, Liu J, Zhang Z, Liu Z, Ou J, Ye M, Zou H (2014) Efficient enrichment of glycopeptides using metal–organic frameworks by hydrophilic interaction chromatography. Analyst 139(19):4987–4993. https://doi.org/10.1039/C4AN00971A

    Article  CAS  PubMed  Google Scholar 

  51. Li Y, Wang H, You X, Ma S, Dong J, Wei Y, Ou J, Ye M (2018) Facile preparation of microporous organic polymers functionalized macroporous hydrophilic resin for selective enrichment of glycopeptides. Anal Chim Acta 1030:96–104. https://doi.org/10.1016/j.aca.2018.05.040

    Article  CAS  PubMed  Google Scholar 

  52. Jiang B, Wu Q, Zhang L, Zhang Y (2017) Preparation and application of silver nanoparticle-functionalized magnetic graphene oxide nanocomposites. Nanoscale 9(4):1607–1615. https://doi.org/10.1039/C6NR09260H

    Article  CAS  PubMed  Google Scholar 

  53. Huang G, Xiong Z, Qin H, Zhu J, Sun Z, Zhang Y, Peng X, Zou H (2014) Synthesis of zwitterionic polymer brushes hybrid silica nanoparticles via controlled polymerization for highly efficient enrichment of glycopeptides. Anal Chim Acta 809:61–68. https://doi.org/10.1016/j.aca.2013.11.049

    Article  CAS  PubMed  Google Scholar 

  54. Zhou J, Liang Y, He X, Chen L, Zhang Y (2017) Dual-functionalized magnetic metal–organic framework for highly specific enrichment of phosphopeptides. ACS Sustain Chem Eng 5(12):11413–11421. https://doi.org/10.1021/acssuschemeng.7b02521

    Article  CAS  Google Scholar 

  55. Stewart II, Thomson T, Figeys D (2001) 18O labeling: a tool for proteomics. Rapid Commun Mass Spectrom 15(24):2456–2465. https://doi.org/10.1002/rcm.525

    Article  CAS  PubMed  Google Scholar 

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Funding

This work is supported by the National Institute of General Medical Sciences and the National Cancer Institute of the National Institutes of Health under Award Numbers R35GM141944 and U01CA185188.

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Authors and Affiliations

  1. Division of Analytical Chemistry, Institute of Chemical Sciences, Bahauddin Zakariya University, Multan, 60800, Pakistan

    Muhammad Salman Sajid, Fahmida Jabeen & Muhammad Najam-ul-Haq

  2. Department of Oncology, Genomics and Epigenomics Shared Resource, Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC, 20057, USA

    Muhammad Salman Sajid & Habtom W. Ressom

  3. Department of Chemistry, The Women University, Kutchery Campus, L.M.Q. Road, Multan, 66000, Pakistan

    Shafaq Saleem

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  1. Muhammad Salman Sajid
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Correspondence to Habtom W. Ressom.

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Sera from subjects recruited at Howard University Hospital (HUH) through IRB Protocol # 2014–0059.

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Additional information includes the Digestion protocols; Sample preparations; Characterizations (IR, and EDX); Selectivity, sensitivity, and reproducibility MALDI-MS spectra; MS/MS spectra and tables.

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Sajid, M.S., Saleem, S., Jabeen, F. et al. Terpolymeric platform with enhanced hydrophilicity via cysteic acid for serum intact glycopeptide analysis. Microchim Acta 189, 277 (2022). https://doi.org/10.1007/s00604-022-05343-0

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