{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,26]],"date-time":"2026-04-26T09:25:23Z","timestamp":1777195523690,"version":"3.51.4"},"reference-count":49,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2022,3,23]],"date-time":"2022-03-23T00:00:00Z","timestamp":1647993600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001659","name":"Deutsche Forschungsgemeinschaft","doi-asserted-by":"publisher","award":["AOBJ655892"],"award-info":[{"award-number":["AOBJ655892"]}],"id":[{"id":"10.13039\/501100001659","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>A novel resemblance-ranking peptide library with 160,000 10-meric peptides was designed to search for selective binders to antibodies. The resemblance-ranking principle enabled the selection of sequences that are most similar to the human peptidome. The library was synthesized with ultra-high-density peptide arrays. As proof of principle, screens for selective binders were performed for the therapeutic anti-CD20 antibody rituximab. Several features in the amino acid composition of antibody-binding peptides were identified. The selective affinity of rituximab increased with an increase in the number of hydrophobic amino acids in a peptide, mainly tryptophan and phenylalanine, while a total charge of the peptide remained relatively small. Peptides with a higher affinity exhibited a lower sum helix propensity. For the 30 strongest peptide binders, a substitutional analysis was performed to determine dissociation constants and the invariant amino acids for binding to rituximab. The strongest selective peptides had a dissociation constant in the hundreds of the nano-molar range. The substitutional analysis revealed a specific hydrophobic epitope for rituximab. To show that conformational binders can, in principle, be detected in array format, cyclic peptide substitutions that are similar to the target of rituximab were investigated. Since the specific binders selected via the resemblance-ranking peptide library were based on the hydrophobic interactions that are widespread in the world of biomolecules, the library can be used to screen for potential linear epitopes that may provide information about the cross-reactivity of antibodies.<\/jats:p>","DOI":"10.3390\/ijms23073515","type":"journal-article","created":{"date-parts":[[2022,3,23]],"date-time":"2022-03-23T22:08:06Z","timestamp":1648073286000},"page":"3515","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Resemblance-Ranking Peptide Library to Screen for Binders to Antibodies on a Peptidomic Scale"],"prefix":"10.3390","volume":"23","author":[{"given":"Felix","family":"Jenne","sequence":"first","affiliation":[{"name":"Institute of Microstructure Technology, Karlsruhe Institute of Technology (KIT), 76344 Eggenstein-Leopoldshafen, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4876-1883","authenticated-orcid":false,"given":"Sergey","family":"Biniaminov","sequence":"additional","affiliation":[{"name":"HS Analysis, Haid-und-Neu-Stra\u00dfe 7, 76131 Karlsruhe, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nathalie","family":"Biniaminov","sequence":"additional","affiliation":[{"name":"HS Analysis, Haid-und-Neu-Stra\u00dfe 7, 76131 Karlsruhe, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Philipp","family":"Marquardt","sequence":"additional","affiliation":[{"name":"HS Analysis, Haid-und-Neu-Stra\u00dfe 7, 76131 Karlsruhe, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Clemens","family":"von Bojni\u010di\u0107-Kninski","sequence":"additional","affiliation":[{"name":"Axxelera, Karl-Floesser-Str. 14, 76189 Karlsruhe, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Roman","family":"Popov","sequence":"additional","affiliation":[{"name":"Axxelera, Karl-Floesser-Str. 14, 76189 Karlsruhe, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2771-712X","authenticated-orcid":false,"given":"Anja","family":"Seckinger","sequence":"additional","affiliation":[{"name":"Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090 Jette, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Dirk","family":"Hose","sequence":"additional","affiliation":[{"name":"Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Laarbeeklaan 103, 1090 Jette, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alexander","family":"Nesterov-Mueller","sequence":"additional","affiliation":[{"name":"Institute of Microstructure Technology, Karlsruhe Institute of Technology (KIT), 76344 Eggenstein-Leopoldshafen, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,3,23]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1138","DOI":"10.1177\/0192623310382559","article-title":"Use of Tissue Cross-reactivity Studies in the Development of Antibody-based Biopharmaceuticals: History, Experience, Methodology, and Future Directions","volume":"38","author":"Leach","year":"2010","journal-title":"Toxicol. Pathol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"11531","DOI":"10.3390\/ijms160511531","article-title":"Optimal Scanning Protocols for Dual-Energy CT Angiography in Peripheral Arterial Stents: An in Vitro Phantom Study","volume":"16","author":"Almutairi","year":"2015","journal-title":"Int. J. Mol. Sci."},{"key":"ref_3","unstructured":"FDA (2022, February 15). Guidances (Drugs), Available online: https:\/\/www.fda.gov\/drugs\/guidance-compliance-regulatory-information\/guidances-drugs."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1509","DOI":"10.1038\/nbt910","article-title":"Analyzing antibody specificity with whole proteome microarrays","volume":"21","author":"Michaud","year":"2003","journal-title":"Nat. Biotechnol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"175","DOI":"10.1038\/nmeth.1178","article-title":"Printing protein arrays from DNA arrays","volume":"5","author":"He","year":"2008","journal-title":"Nat. Methods"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"86","DOI":"10.1126\/science.1097639","article-title":"Self-Assembling Protein Microarrays","volume":"305","author":"Ramachandran","year":"2004","journal-title":"Science"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1658","DOI":"10.1074\/mcp.T600024-MCP200","article-title":"Generation of High Density Protein Microarrays by Cell-free in Situ Expression of Unpurified PCR Products","volume":"5","author":"Angenendt","year":"2006","journal-title":"Mol. Cell. Proteom."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"666","DOI":"10.1038\/nsmb.1842","article-title":"Post-translational modifications in signal integration","volume":"17","author":"Deribe","year":"2010","journal-title":"Nat. Struct. Mol. Biol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"231","DOI":"10.2144\/0000113922","article-title":"Real-time protein unfolding: A method for determining the kinetics of native protein denaturation using a quantitative real-time thermocycler","volume":"53","author":"Biggar","year":"2012","journal-title":"BioTechniques"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"535","DOI":"10.1038\/nbt.1856","article-title":"Autoantigen discovery with a synthetic human peptidome","volume":"29","author":"Larman","year":"2011","journal-title":"Nat. Biotechnol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"622","DOI":"10.1016\/j.ab.2011.11.005","article-title":"Phage display screening without repetitious selection rounds","volume":"421","author":"Hoen","year":"2012","journal-title":"Anal. Biochem."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"790","DOI":"10.3390\/molecules16010790","article-title":"Phage Display: Selecting Straws Instead of a Needle from a Haystack","volume":"16","author":"Vodnik","year":"2011","journal-title":"Molecules"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1016\/j.jim.2017.01.012","article-title":"Single amino acid fingerprinting of the human antibody repertoire with high density peptide arrays","volume":"443","author":"Weber","year":"2017","journal-title":"J. Immunol. Methods"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1078","DOI":"10.1002\/elsc.201700062","article-title":"Antibody fingerprints in lyme disease deciphered with high density peptide arrays","volume":"17","author":"Weber","year":"2017","journal-title":"Eng. Life Sci."},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Palermo, A., and Nesterov-Mueller, A. (2019). Serological Number for Characterization of Circulating Antibodies. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20030604"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"463","DOI":"10.1021\/cc100009g","article-title":"Automated Maskless Photolithography System for Peptide Microarray Synthesis on a Chip","volume":"12","author":"Shin","year":"2010","journal-title":"J. Comb. Chem."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1790","DOI":"10.1074\/mcp.M112.020800","article-title":"High-resolution Mapping of Linear Antibody Epitopes Using Ultrahigh-density Peptide Microarrays","volume":"11","author":"Buus","year":"2012","journal-title":"Mol. Cell. Proteom."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"767","DOI":"10.1126\/science.1990438","article-title":"Light-Directed, Spatially Addressable Parallel Chemical Synthesis","volume":"251","author":"Fodor","year":"1991","journal-title":"Science"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"4785","DOI":"10.1038\/ncomms5785","article-title":"Scalable high-density peptide arrays for comprehensive health monitoring","volume":"5","author":"Legutki","year":"2014","journal-title":"Nat. Commun."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"7359","DOI":"10.1038\/sj.onc.1206939","article-title":"Rituximab (monoclonal anti-CD20 antibody): Mechanisms of action and resistance","volume":"22","author":"Smith","year":"2003","journal-title":"Oncogene"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"4075","DOI":"10.1529\/biophysj.104.049288","article-title":"Helical Packing Patterns in Membrane and Soluble Proteins","volume":"87","author":"Gimpelev","year":"2004","journal-title":"Biophys. J."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"927","DOI":"10.1016\/j.ekir.2020.03.009","article-title":"In situ Visualization of C3\/C5 Convertases to Differentiate Complement Activation","volume":"5","author":"Person","year":"2020","journal-title":"Kidney Int. Rep."},{"key":"ref_23","unstructured":"(2022, February 15). UniPro. Available online: http:\/\/www.uniprot.org."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"13036","DOI":"10.1038\/s41598-019-49539-6","article-title":"Efficient partition of integer optimization problems with one-hot encoding","volume":"9","author":"Okada","year":"2019","journal-title":"Sci. Rep."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"43","DOI":"10.1016\/j.critrevonc.2006.09.004","article-title":"Pharmacokinetics of rituximab and its clinical use: Thought for the best use?","volume":"62","author":"Cartron","year":"2007","journal-title":"Crit. Rev. Oncol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"22","DOI":"10.2174\/157488708783330495","article-title":"Pharmacokinetic properties of rituximab","volume":"3","author":"Rodriguez","year":"2008","journal-title":"Rev. Recent Clin. Trials"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"995","DOI":"10.1023\/A:1008416911099","article-title":"Association of serum Rituximab (IDEC\u2013C2B8) concentration and anti-tumor response in the treatment of recurrent low-grade or follicular non-Hodgkin\u2019s lymphoma","volume":"9","author":"Berinstein","year":"1998","journal-title":"Ann. Oncol."},{"key":"ref_28","first-page":"369","article-title":"Hydrophobic Parameters-Pi of Amino-Acid Side-Chains from the Partitioning of N-Acetyl-Amino-Acid Amides","volume":"18","author":"Fauchere","year":"1983","journal-title":"Eur. J. Med. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"509","DOI":"10.1038\/nchem.1062","article-title":"Contemporary strategies for peptide macrocyclization","volume":"3","author":"White","year":"2011","journal-title":"Nat. Chem."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2095","DOI":"10.1021\/acs.orglett.9b00378","article-title":"Synthesis of Constrained Tetracyclic Peptides by Consecutive CEPS, CLIPS, and Oxime Ligation","volume":"21","author":"Streefkerk","year":"2019","journal-title":"Org. Lett."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"422","DOI":"10.1016\/S0006-3495(98)77529-0","article-title":"A Helix Propensity Scale Based on Experimental Studies of Peptides and Proteins","volume":"75","author":"Pace","year":"1998","journal-title":"Biophys. J."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"168","DOI":"10.1038\/nature04177","article-title":"A quantitative protein interaction network for the ErbB receptors using protein microarrays","volume":"439","author":"Jones","year":"2006","journal-title":"Nature"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"6865","DOI":"10.1021\/acs.analchem.1c00960","article-title":"Determination of the Rituximab Binding Site to the CD20 Epitope Using SPOT Synthesis and Surface Plasmon Resonance Analyses","volume":"93","author":"Bar","year":"2021","journal-title":"Anal. Chem."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"6776","DOI":"10.1073\/pnas.1700818114","article-title":"Weak protein-protein interactions in live cells are quantified by cell-volume modulation","volume":"114","author":"Sukenik","year":"2017","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"3724","DOI":"10.1038\/ncomms4724","article-title":"Protein conformational dynamics dictate the binding affinity for a ligand","volume":"5","author":"Seo","year":"2014","journal-title":"Nat. Commun."},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Popov, R., Shankara, G.K., von Bojni\u010di\u0107-Kninski, C., and Nesterov-Mueller, A. (2019). Renaissance Distribution for Statistically Failed Experiments. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20133250"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1975","DOI":"10.1182\/blood-2006-04-014639","article-title":"The epitope recognized by rituximab","volume":"108","author":"Binder","year":"2006","journal-title":"Blood"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"15073","DOI":"10.1074\/jbc.M701654200","article-title":"Structural Basis for Recognition of CD20 by Therapeutic Antibody Rituximab","volume":"282","author":"Du","year":"2007","journal-title":"J. Biol. Chem."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"8357","DOI":"10.1016\/S0040-4039(98)01843-7","article-title":"Efficient synthesis of thioether-based cyclic peptide libraries","volume":"39","author":"Roberts","year":"1998","journal-title":"Tetrahedron Lett."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"3256","DOI":"10.1182\/blood.V99.9.3256","article-title":"Alanine-170 and proline-172 are critical determinants for extracellular CD20 epitopes; heterogeneity in the fine specificity of CD20 monoclonal antibodies is defined by additional requirements imposed by both amino acid sequence and quaternary structure","volume":"99","author":"Polyak","year":"2002","journal-title":"Blood"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1070","DOI":"10.1182\/blood-2005-04-1769","article-title":"Generation of biologically active linear and cyclic peptides has revealed a unique fine specificity of rituximab and its possible cross-reactivity with acid sphingomyelinase-like phosphodiesterase 3b precursor","volume":"107","author":"Perosa","year":"2006","journal-title":"Blood"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"85ps21","DOI":"10.1126\/scitranslmed.3002429","article-title":"Rituximab\u2019s New Therapeutic Target: The Podocyte Actin Cytoskeleton","volume":"3","author":"Chan","year":"2011","journal-title":"Sci. Transl. Med."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"85ra46","DOI":"10.1126\/scitranslmed.3002231","article-title":"Rituximab Targets Podocytes in Recurrent Focal Segmental Glomerulosclerosis","volume":"3","author":"Fornoni","year":"2011","journal-title":"Sci. Transl. Med."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"534","DOI":"10.1016\/j.drudis.2007.05.004","article-title":"Water, water everywhere\u2014Except where it matters?","volume":"12","author":"Homans","year":"2007","journal-title":"Drug Discov. Today"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"973","DOI":"10.1016\/S1074-5521(96)90164-7","article-title":"Just add water! The effect of water on the specificity of protein-ligand binding sites and its potential application to drug design","volume":"3","author":"Ladbury","year":"1996","journal-title":"Chem. Biol."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s41467-018-05769-2","article-title":"Intriguing role of water in protein-ligand binding studied by neutron crystallography on trypsin complexes","volume":"9","author":"Schiebel","year":"2018","journal-title":"Nat. Commun."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"569","DOI":"10.1038\/s41467-022-28164-4","article-title":"Decisive role of water and protein dynamics in residence time of p38\u03b1 MAP kinase inhibitors","volume":"13","author":"Pantsar","year":"2022","journal-title":"Nat. Commun."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"396","DOI":"10.1016\/j.ccell.2017.02.002","article-title":"Target Expression, Generation, Preclinical Activity, and Pharmacokinetics of the BCMA-T Cell Bispecific Antibody EM801 for Multiple Myeloma Treatment","volume":"31","author":"Seckinger","year":"2017","journal-title":"Cancer Cell"},{"key":"ref_49","unstructured":"(2022, February 15). AXXELERA UG. Available online: https:\/\/axxelera.com\/."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/23\/7\/3515\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T22:42:00Z","timestamp":1760136120000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/23\/7\/3515"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,3,23]]},"references-count":49,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2022,4]]}},"alternative-id":["ijms23073515"],"URL":"https:\/\/doi.org\/10.3390\/ijms23073515","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,3,23]]}}}