Antibodies represent an essential component of humoral immunity; therefore their study is important for molecular biology and medicine. The unique property of antibodies to specifically recognize and bind a certain molecular target (an antigen) determines their widespread application in treatment and diagnostics of diseases, as well as in laboratory and biotechnological practices. High specificity and affinity of antibodies is determined by the presence of primary structure variable regions, which are not encoded in the human genome and are unique for each antibody-producing B cell clone. Hence, there is little or no information about amino acid sequences of the variable regions in the databases. This differs identification of antibody primary structure from most of the proteomic studies because it requires either B cell genome sequencing or de novo amino acid sequencing of the antibody. The present review demonstrates some examples of proteomic and proteogenomic approaches and the methodological arsenal that proteomics can offer for studying antibodies, in particular, for identification of primary structure, evaluation of posttranslational modifications and application of bioinformatics tools for their decoding.
Download PDF:
Keywords: antibodies, mass-spectrometry, de novo antibody sequencing, antibody repertoire
Kazieva, L. Sh., Farafonova, T. E., Zgoda, V. G. (2023). Antibody proteomics. Biomeditsinskaya Khimiya, 69(1), 5-18.
References
Yman V., Tuju J., White M.T., Kamuyu G., Mwai K., Kibinge N., Asghar M., Sundling C., Sondén K., Murungi L. et al. (2022) Distinct kinetics of antibodies to 111 Plasmodium falciparum proteins identifies markers of recent malaria exposure. Nat. Commun., 13(1), 331-331. CrossRef Scholar google search
Monroy-Iglesias M.J., Crescioli S., Beckmann K., Le N., Karagiannis S.N., van Hemelrijck M., Santaolalla A. (2022) Antibodies as biomarkers for cancer risk: A systematic review. Clin. Exper. Immunol., 209(1), 46-63. CrossRef Scholar google search
Satoh M., Ceribelli A., Hasegawa T., Tanaka S. (2022) Clinical significance of antinucleolar antibodies: Biomarkers for autoimmune diseases, malignancies, and others. Clin. Rev. Allergy Immunol., 63(2), 210-239. CrossRef Scholar google search
Alsaed O.S., Alamlih L.I., Al-Radideh O., Chandra P., Alemadi S., Al-Allaf A.-W. (2021) Clinical utility of ANA-ELISA vs ANA-immunofluorescence in connective tissue diseases. Sci. Rep., 11(1), 8229-8229. CrossRef Scholar google search
Musharova O., Medvedeva S., Klimuk E., Guzman N.M., Titova D., Zgoda V., Shiriaeva A., Semenova E., Severinov K., Savitskaya E. (2021) Prespacers formed during primed adaptation associate with the Cas1-Cas2 adaptation complex and the Cas3 interference nuclease-helicase. Proc. Nat. Acad. Sci. USA, 118(22), e2021291118. CrossRef Scholar google search
Neubert H., Shuford C.M., Olah T.V., Garofolo F., Schultz G.A., Jones B.R., Amaravadi L., Laterza O.F., Xu K., Ackermann B.L. (2020) Protein biomarker quantification by immunoaffinity liquid chromatography-tandem mass spectrometry: Current state and future vision. Clinical Chemistry, 66(2), 282-301. CrossRef Scholar google search
Brummelman J., Haftmann C., Núñez N.G., Alvisi G., Mazza E.M.C., Becher B., Lugli E. (2019) Development, application and computational analysis of high-dimensional fluorescent antibody panels for single-cell flow cytometry. Nature Protocols, 14(7), 1946-1969. CrossRef Scholar google search
David M.P.C., Asprer J.J.T., Ibana J.S.A., Concepcion G.P., Padlan E.A. (2007) A study of the structural correlates of affinity maturation: antibody affinity as a function of chemical interactions, structural plasticity and stability. Mol. Immunol., 44(6), 1342-1351. CrossRef Scholar google search
Stanfield R.L., Wilson I.A. (2014) Antibody structure. Microbiology Spectrum, 2(2), DOI: 10.1128/microbiolspec.AID-0012-2013. CrossRef Scholar google search
Snapkov I., Chernigovskaya M., Sinitcyn P., Lê Quý K., Nyman T.A., Greiff V. (2022) Progress and challenges in mass spectrometry-based analysis of antibody repertoires. Trends Biotechnol., 40(4), 463-481. CrossRef Scholar google search
Greiff V., Miho E., Menzel U., Reddy S.T. (2015) Bioinformatic and statistical analysis of adaptive immune repertoires. Trends Immunol., 36(11), 738-749. CrossRef Scholar google search
Xu J.L., Davis M.M. (2000) Diversity in the CDR3 region of V(H) is sufficient for most antibody specificities. Immunity, 13(1), 37-45. CrossRef Scholar google search
Khass M., Vale A.M., Burrows P.D., Schroeder H.W. (2018) The sequences encoded by immunoglobulin diversity (DH) gene segments play key roles in controlling B-cell development, antigen-binding site diversity, and antibody production. Immunol. Rev., 284(1), 106-119. CrossRef Scholar google search
Robinson S.A., Raybould M.I.J., Schneider C., Wong W.K., Marks C., Deane C.M. (2021) Epitope profiling using computational structural modelling demonstrated on coronavirus-binding antibodies. PLoS Comput. Biol., 17(12), e1009675. CrossRef Scholar google search
Lu R.-M., Hwang Y.-C., Liu I.-J., Lee C.-C., Tsai H.-Z., Li H.-J., Wu H.-C. (2020) Development of therapeutic antibodies for the treatment of diseases. J. Biomed. Sci., 27(1), 1-1. CrossRef Scholar google search
Awwad S., Angkawinitwong U. (2018) Overview of antibody drug delivery. Pharmaceutics, 10(3), 83-83. CrossRef Scholar google search
Frejd F.Y., Kim K.-T. (2017) Affibody molecules as engineered protein drugs. Exper. Mol. Med., 49(3), e306-e306. CrossRef Scholar google search
Brasino M., Roy S., Erbse A.H., He L., Mao C., Park W., Cha J.N., Goodwin A.P. (2018) Anti-EGFR affibodies with site-specific photo-cross-linker incorporation show both directed target-specific photoconjugation and increased retention in tumors. J. Am. Chem. Soc., 140(37), 11820-11828. CrossRef Scholar google search
Altunay B., Morgenroth A., Beheshti M., Vogg A., Wong N.C.L., Ting H.H., Biersack H.-J., Stickeler E., Mottaghy F.M. (2021) HER2-directed antibodies, affibodies and nanobodies as drug-delivery vehicles in breast cancer with a specific focus on radioimmunotherapy and radioimmunoimaging. Eur. J. Nucl. Med. Mol. Imaging, 48(5), 1371-1389. CrossRef Scholar google search
Lee S.C., Ma J.S.Y., Kim M.S., Laborda E., Choi S.-H., Hampton E.N., Yun H., Nunez V., Muldong M.T., Wu C.N., Ma W. et al. (2021) A PSMA-targeted bispecific antibody for prostate cancer driven by a small-molecule targeting ligand. Science Advances, 7(33), eabi8193. CrossRef Scholar google search
Oldham R.K., Dillman R.O. (2008) Monoclonal antibodies in cancer therapy: 25 years of progress. J. Clin. Oncol., 26(11), 1774-1777. CrossRef Scholar google search
Hafeez U., Gan H.K., Scott A.M. (2018) Monoclonal antibodies as immunomodulatory therapy against cancer and autoimmune diseases. Curr. Opin. Pharmacol., 41, 114-121. CrossRef Scholar google search
Melero I., Hervas-Stubbs S., Glennie M., Pardoll D.M., Chen L. (2007) Immunostimulatory monoclonal antibodies for cancer therapy. Nature Reviews. Cancer, 7(2), 95-106. CrossRef Scholar google search
Fujiwara S., Wada H., Kawada J., Kawabata R., Takahashi T., Fujita J., Hirao T., Shibata K., Makari Y., Iijima S. et al. (2013) NY-ESO-1 antibody as a novel tumour marker of gastric cancer. Br. J. Cancer, 108(5), 1119-1125. CrossRef Scholar google search
Lee Z.J.O., Eslick G.D., Edirimanne S. (2020) Investigating antithyroglobulin antibody as a prognostic marker for differentiated thyroid cancer: A meta-analysis and systematic review. Thyroid, 30(11), 1601-1612. CrossRef Scholar google search
Bogdanos D.P., Gkoutzourelas A., Papadopoulos V., Liaskos C., Patrikiou E., Tsigalou C., Saratziotis A., Hajiioannou J., Scheper T., Meyer W., Sakkas L.I., Papandreou C. (2021) Anti-Ro52 antibody is highly prevalent and a marker of better prognosis in patients with ovarian cancer. Clin. Chim. Acta, 521, 199-205. CrossRef Scholar google search
Scherer H.U., Huizinga T.W.J., Krönke G., Schett G., Toes R.E.M. (2018) The B cell response to citrullinated antigens in the development of rheumatoid arthritis. Nature Reviews. Rheumatology, 14(3), 157-169. CrossRef Scholar google search
Darrah E., Giles J.T., Ols M.L., Bull H.G., Andrade F., Rosen A. (2013) Erosive rheumatoid arthritis is associated with antibodies that activate PAD4 by increasing calcium sensitivity. Sci. Transl. Med., 5(186), 186ra65. CrossRef Scholar google search
Zhou X., Motta F., Selmi C., Ridgway W.M., Gershwin M.E., Zhang W. (2021) Antibody glycosylation in autoimmune diseases. Autoimmun. Rev., 20(5), 102804-102804. CrossRef Scholar google search
Fechtner S., Berens H., Bemis E., Johnson R.L., Guthridge C.J., Carlson N.E., Demoruelle M.K., Harley J.B., Edison J.D., Norris J.A. et al. (2022) Antibody responses to Epstein-Barr virus in the preclinical period of rheumatoid arthritis suggest the presence of increased viral reactivation cycles. Arthritis Rheumatology, 74(4), 597-603. CrossRef Scholar google search
Lu L.L., Chung A.W., Rosebrock T.R., Ghebremichael M., Yu W.H., Grace P.S., Schoen M.K., Tafesse F., Martin C., Leung V. et al. (2016) A functional role for antibodies in tuberculosis. Cell, 167(2), 433-443.e14. CrossRef Scholar google search
Alter G., Ottenhoff T.H.M., Joosten S.A. (2018) Antibody glycosylation in inflammation, disease and vaccination. Semin. Immunol., 39, 102-110. CrossRef Scholar google search
Offersen R., Yu W.-H., Scully E.P., Julg B., Euler Z., Sadanand S., Garcia-Dominguez D., Zheng L., Rasmussen T.A., Jennewein M.F. et al. (2020) HIV antibody Fc N-linked glycosylation is associated with viral rebound. Cell Reports, 33(11), 108502-108502. CrossRef Scholar google search
Bournazos S., Vo H.T.M., Duong V., Auerswald H., Ly S., Sakuntabhai A., Dussart P., Cantaert T., Ravetch J.V. (2021) Antibody fucosylation predicts disease severity in secondary dengue infection. Science, 372(6546), 1102-1105. CrossRef Scholar google search
Toby T.K., Fornelli L., Srzentić K., de Hart C.J., Levitsky J., Friedewald J., Kelleher N.L. (2019) A comprehensive pipeline for translational top-down proteomics from a single blood draw. Nature Protocols, 14(1), 119-152. CrossRef Scholar google search
Hirosawa M., Hoshida M., Ishikawa M., Toya T. (1993) MASCOT: Multiple alignment system for protein sequences based on three-way dynamic programming. Bioinformatics, 9(2), 161-167. CrossRef Scholar google search
Tyanova S., Temu T., Cox J. (2016) The MaxQuant computational platform for mass spectrometry-based shotgun proteomics. Nature Protocols, 11(12), 2301-2319. CrossRef Scholar google search
Barsnes H., Vaudel M. (2018) SearchGUI: A highly adaptable common interface for proteomics search and de novo engines. J. Proteome Res., 17(7), 2552-2555. CrossRef Scholar google search
Vaudel M., Burkhart J.M., Zahedi R.P., Oveland E., Berven F.S., Sickmann A., Martens L., Barsnes H. (2015) PeptideShaker enables reanalysis of MS-derived proteomics data sets. Nature Biotechnology, 33(1), 22-24. CrossRef Scholar google search
Zhang J., Xin L., Shan B., Chen W., Xie M., Yuen D., Zhang W., Zhang Z., Lajoie G.A., Ma B. (2012) PEAKS DB: De novo sequencing assisted database search for sensitive and accurate peptide identification. Mol. Cell. Proteomics, 11(4), M111.010587-M111.010587. CrossRef Scholar google search
Frank A., Pevzner P. (2005) PepNovo: De novo peptide sequencing via probabilistic network modeling. Anal. Chem., 77(4), 964-973. CrossRef Scholar google search
Jeong K., Kim S., Pevzner P.A. (2013) UniNovo: A universal tool for de novo peptide sequencing. Bioinformatics, 29(16), 1953-1962. CrossRef Scholar google search
Sen K.I., Tang W.H., Nayak S., Kil Y.J., Bern M., Ozoglu B., Ueberheide B., Davis D., Becker C. (2017) Automated antibody de novo sequencing and its utility in biopharmaceutical discovery. J. Am. Soc. Mass Spectrom., 28(5), 803-810. CrossRef Scholar google search
Zhang Y., Fonslow B.R., Shan B., Baek M.-C., Yates J.R. (2013) Protein analysis by shotgun/bottom-up proteomics. Chem. Rev., 113(4), 2343-2394. CrossRef Scholar google search
Peng W., Pronker M.F., Snijder J. (2021) Mass spectrometrybased de novo sequencing of monoclonal antibodies using multiple proteases and a dual fragmentation scheme. J. Proteome Res., 20, 3559-3566. CrossRef Scholar google search
Cristobal A., Marino F., Post H., van den Toorn H.W.P., Mohammed S., Heck A.J.R. (2017) Toward an optimized workflow for middle-down proteomics. Anal. Chem., 89(6), 3318-3325. CrossRef Scholar google search
Fornelli L., Srzentić K., Huguet R., Mullen C., Sharma S., Zabrouskov V., Fellers R.T., Durbin K.R., Compton P.D., Kelleher N.L. (2018) Accurate sequence analysis of a monoclonal antibody by top-down and middle-down orbitrap mass spectrometry applying multiple ion activation techniques. Anal. Chem., 90(14), 8421-8429. CrossRef Scholar google search
Xiao Y., Vecchi M.M., Wen D. (2016) Distinguishing between leucine and isoleucine by integrated LC-MS analysis using an orbitrap fusion mass spectrometer. Anal. Chem., 88(21), 10757-10766. CrossRef Scholar google search
Mao Y., Valeja S.G., Rouse J.C., Hendrickson C.L., Marshall A.G. (2013) Top-down structural analysis of an intact monoclonal antibody by electron capture dissociation-fourier transform ion cyclotron resonancemass spectrometry. Anal. Chem., 85(9), 4239-4246. CrossRef Scholar google search
Fornelli L., Ayoub D., Aizikov K., Beck A., Tsybin Y.O. (2014) Middle-down analysis of monoclonal antibodies with electron transfer dissociation orbitrap fourier transform mass spectrometry. Anal. Chem., 86(6), 3005-3012. CrossRef Scholar google search
Wang Z., Liu X., Muther J., James J.A., Smith K., Wu S. (2019) Top-down mass spectrometry analysis of human serum autoantibody antigen-binding fragments. Sci. Rep., 9(1), 2345. CrossRef Scholar google search
Rosati S., Yang Y., Barendregt A., Heck A.J.R. (2014) Detailed mass analysis of structural heterogeneity in monoclonal antibodies using native mass spectrometry. Nature Protocols, 9(4), 967-976. CrossRef Scholar google search
Cheng J., Wang L., Rive C.M., Holt R.A., Morin G.B., Chen D.D.Y. (2020) Complementary methods for de novo monoclonal antibody sequencing to achieve complete sequence coverage. J. Proteome Res., 19(7), 2700-2707. CrossRef Scholar google search
Wheatley A.K., Juno J.A., Wang J.J., Selva K.J., Reynaldi A., Tan H.-X., Lee W.S., Wragg K.M., Kelly H.G., Esterbauer R., Davis S.K. et al. (2021) Evolution of immune responses to SARS-CoV-2 in mild-moderate COVID-19. Nat. Commun., 12(1), 1162-1162. CrossRef Scholar google search
Lavinder J.J., Wine Y., Giesecke C., Ippolito G.C., Horton A.P., Lungu O.I., Hoi K.H., de Kosky B.J., Murrin E.M., Wirth M.M. et al. (2014) Identification and characterization of the constituent human serum antibodies elicited by vaccination. Proc. Nat. Acad. Sci. USA, 111(6), 2259-2264. CrossRef Scholar google search
Guthals A., Gan Y., Murray L., Chen Y., Stinson J., Nakamura G., Lill J.R., Sandoval W., Bandeira N. (2017) De novo MS/MS sequencing of native human antibodies. J. Proteome Res., 16(1), 45-54. CrossRef Scholar google search
Lee J., Paparoditis P., Horton A.P., Frühwirth A., McDaniel J.R., Jung J., Boutz D.R., Hussein D.A., Tanno Y., Pappas L. et al. (2019) Persistent antibody clonotypes dominate the serum response to influenza over multiple years and repeated vaccinations. Cell Host Microbe, 25(3), 367-376.e5. CrossRef Scholar google search
Bondt A., Dingess K.A., Hoek M., van Rijswijck D.M.H., Heck A.J.R. (2021) A direct MS-based approach to profile human milk secretory immunoglobulin A (IgA1) reveals donor-specific clonal repertoires with high longitudinal stability. Front. Immunol., 12, 789748. CrossRef Scholar google search
Wine Y., Boutz D.R., Lavinder J.J., Miklos A.E., Hughes R.A., Hoi K.H., Jung S.T., Horton A.P., Murrin E.M., Ellington A.D. et al. (2013) Molecular deconvolution of the monoclonal antibodies that comprise the polyclonal serum response. Proc. Nat. Acad. Sci., 110(8), 2993-2998. CrossRef Scholar google search
Shaw J.B., Liu W., Vasil'ev Y.V., Bracken C.C., Malhan N., Guthals A., Beckman J.S., Voinov V.G. (2020) Direct determination of antibody chain pairing by top-down and middle-down mass spectrometry using electron capture dissociation and ultraviolet photodissociation. Anal. Chem., 92(1), 766-773. CrossRef Scholar google search
Srzentić K., Nagornov K.O., Fornelli L., Lobas A.A., Ayoub D., Kozhinov A.N., Gasilova N., Menin L., Beck A., Gorshkov M.V., Aizikov K., Tsybin Y.O. (2018) Multiplexed middle-down mass spectrometry as a method for revealing light and heavy chain connectivity in a monoclonal antibody. Anal. Chem., 90(21), 12527-12535. CrossRef Scholar google search
Boutz D.R., Horton A.P., Wine Y., Lavinder J.J., Georgiou G., Marcotte E.M. (2014) Proteomic identification of monoclonal antibodies from serum. Anal. Chem., 86(10), 4758-4766. CrossRef Scholar google search
Lundström S.L., Zhang B., Rutishauser D., Aarsland D., Zubarev R.A. (2017) SpotLight proteomics: Uncovering the hidden blood proteome improves diagnostic power of proteomics. Sci. Rep., 7(1), 41929-41929. CrossRef Scholar google search
Zhang B., Pirmoradian M., Chernobrovkin A., Zubarev R.A. (2014) DeMix workflow for efficient identification of cofragmented peptides in high resolution data-dependent tandem mass spectrometry. Mol. Cell. Proteomics, 13(11), 3211-3223. CrossRef Scholar google search
Bern M., Kil Y.J., Becker C. (2012) Byonic: Advanced peptide and protein identification software. Curr. Protoc. Bioinformatics, 40(1), 13.20.1-13.20.14. CrossRef Scholar google search
Schulte D., Peng W., Snijder J. (2022) Template-based assembly of proteomic short reads for de novo antibody sequencing and repertoire profiling. Anal. Chem., 94(29), 10391-10399. CrossRef Scholar google search
