Research and Development of High-Affinity Binding Proteins

Research and Development of High-Affinity Binding Proteins

RNDr. Petr Malý, CSc.

RNDr. Petr Malý, CSc. — Project head

The Institute of Biotechnology of the Czech Academy of Sciences

About us

The project systematically studies small protein ligands and actively develops methods of their engineering by means of molecular and structural biology and also builds know-how for construction of proteins with binding properties. Protein ligands binding to cytokines and their receptors, interferon γ, proteins of interleukins 23, 12, 17, and of their receptors are designed and prepared.


  • Design and preparation of small protein ligands binding target proteins of biotechnological, diagnostic, and/or medical importance with high affinity and specificity
  • Commercial preparation of custom-made proteins

Artificial binding proteins derived from small protein scaffolds represent a valuable alternative to commonly used antibodies. Novel binders with engineered affinity, high specificity or designed inhibitory function attract attention as key components for the development of novel biosensing devices, in vivo diagnostics and next-generation therapeutics. Small, robust, and soluble proteins with high thermal and hydrodynamic stability and without disulphide bonds are amenable to rational improvement and can be easily modified by gene-fusion approaches.


In our laboratory, which operates since 2008, we focus on engineering novel binders raised against human cytokines, their receptors or tumour markers. Recently we have established a model of three-helix bundle of albumin-binding domain (ABD) of streptococcal protein G as a master scaffold for the generation of high-complex combinatorial libraries. Using randomization of 11 pre-determined amino acid residues we engineered a library of a theoretical complexity of 1014 protein variants which was successfully used, in combination with ribosome display selection, for the generation of unique binders of human interferon gamma (hIFNg) with KD in the nanomolar range (Ahmad et al 2012). High-affinity binders of hIFNg were then used as key components for the construction of a novel SPR biosensor with the improved sensitivity for detection of hIFNg in diluted blood plasma (Šípová et al. 2012).





Kosztyu, P., Kuchar, M., Cerny, J., Barkocziova, L., Maly, M., Petrokova, H., Czernekova, L., Liskova, V., Raskova Kafkova, L., Knotigova, P., Masek, J., Turanek, J., Maly, P., Raska M. Proteins mimicking epitope of HIV-1 virus neutralizing antibody induce virus-neutralizing sera in mice. EBioMedicine, 47: 247–256, 2019. doi: 10.1016/j.ebiom.2019.07.015. ISSN: 2352-3964.

Plavec, T.V., Kuchar, M., Benko, A., Liskova, V., Cerny, J., Berlec, A., Maly, P. Engineered Lactococcus lactis Secreting IL-23 Receptor-Targeted REX Protein Blockers for Modulation of IL-23/Th17-Mediated Inflammation. Microorganisms, 7(5): 152, 2019. doi: 10.3390/microorganisms7050152. EISSN 2076-2607.


Hlavnickova, M., Kuchar, M., Osicka, R., Vankova, L., Petrokova, H., Maly, M., Cerny, J., Arenberger, P., Maly, P. ABD-Derived Protein Blockers of Human IL-17 Receptor A as Non-IgG Alternatives for Modulation of IL-17-Dependent Pro-Inflammatory Axis. International Journal of Molecular Sciences, 19(10): 3089, 2018. doi: 10.3390/ijms19103089. eISSN: 1422-0067.

Skrlec, K., Zadravec, P., Hlavnickova, M., Kuchar, M., Vankova, L., Petrokova, H., Krizova, L., Cerny, J., Berlec, A., Maly, P. p19-Targeting ILP Protein Blockers of IL-23/Th-17 Pro-Inflammatory Axis Displayed on Engineered Bacteria of Food Origin. International Journal of Molecular Sciences, 19(7): 1933, 2018. doi: 10.3390/ijms19071933. ISSN 1422-0067.

Semeradtova, A., Stofik, M., Vankova, L., Maly, P., Stanek, O., Maly, J. Optical microchips based on high-affinity recombinant protein binders - Human serum albumin detection in urine. Sensors and Actuators B: Chemical, 272: 441-447, 2018. doi: 10.1016/j.snb.2018.05.180. ISSN: 0925-4005.


Krizova, L., Kuchar, M., Petrokova, H., Osicka, R., Hlavnickova, M., Pelak, O., Cerny, J., Kalina, T., Maly, P. p19-targeted ABD-derived protein variants inhibit IL-23 binding and exert suppressive control over IL-23-stimulated expansion of primary human IL-17+ T-cells. Autoimmunity, 50: 102-113, 2017. doi: 10.1080/08916934.2016.1272598. ISSN 0891-6934.


Maly, J., Stanek, O., Frolik, J., Maly, M., Ennen, F., Appelhans, D., Semeradtova, A., Wrobel, D., Stofik, M., Knapova, T., Kuchar, M., Stastna, L., Cermak, J., Sebo, P., Maly, P. Biocompatible Size-Defined Dendrimer–Albumin Binding Protein Hybrid Materials as a Versatile Platform for Biomedical Applications. Macromolecular Bioscience, 16(4): 553-566, 2016. doi: 10.1002/mabi.201500332. eISSN 1616-5195.

Zadravec, P., Mareckova, L., Petrokova, H., Hodnik, V., Nanut, M. P., Anderluh, G., Strukelj, B., Maly, P., Berlec, A. Development of Recombinant Lactococcus lactis Displaying Albumin-Binding Domain Variants against Shiga Toxin 1 B Subunit. PLoS ONE 11(9): e0162625, 2016. doi: 10.1371/journal.pone.0162625. ISSN 1932-6203.


Marečková L, Petroková H, Osička R, Kuchař M, Malý P. Novel binders derived from an albumin-binding domain scaffold targeting human prostate secretory protein 94 (PSP94). Protein Cell. 2015 Oct;6(10):774-9.

Mareckova, L., Petrokova, H., Osicka, R., Kuchar, M., Maly, P. Novel binders derived from an albumin binding domain scaffold targeting human prostate secretory protein 94 (PSP94)v. Protein & Cell, 6(10): 774-779, 2015. doi: 10.1007/s13238-015-0194-9. ISSN 1674-8018.


Milan Kuchar, Lucie Vankova, Hana Petrokova, Jiri Cerny, Radim Osicka, Ondrej Pelak, Hana Sipova, Bohdan Schneider, Jiri Homola, Peter Sebo, Tomas Kalina & Petr Maly: Human interleukin-23 receptor antagonists derived from an albumin-binding domain scaffold inhibit IL-23-dependent ex vivo expansion of IL-17-producing T-cells. Proteins 82, 975-989 (2014). doi:10.1002/prot.24472.


Kuchař M, Vaňková L, Petroková H, Cerný J, Osička R, Pelák O, Sípová H, Schneider B, Homola J, Sebo P, Kalina T, Malý P. Human interleukin-23 receptor antagonists derived from an albumin-binding domain scaffold inhibit IL-23-dependent ex vivo expansion of IL-17-producing T-cells. Proteins. 2014 Jun;82(6):975-89. doi: 10.1002/prot.24472. Epub 2013 Nov 23.


Ahmad JN, Li J, Biedermannová L, Kuchař M, Sípová H, Semerádtová A, Cerný J, Petroková H, Mikulecký P, Polínek J, Staněk O, Vondrášek J, Homola J, Malý J, Osička R, Sebo P, Malý P. Novel high-affinity binders of human interferon gamma derived from albumin-binding domain of protein G. Proteins. 2012 Mar;80(3):774-89.

Šípová H, Ševců V, Kuchař M, Ahmad JN, Mikulecký P, Osička R, Malý P, Homola J. Surface plasmon resonance biosensor based on engineered proteins for direct detection of interferon-gamma in diluted blood plasma. Sensors and Actuators B, 2012,


Krejcirikova V, Pachl P., Fabry M, Maly P, Rezacova P, Brynda J. Structure of the mouse galectin-4 N-terminal carbohydrate-recognition domain reveals the mechanism of oligosaccharide recognition. Acta Crystallographica Section D, Biological Crystallography, 2011, D67, 204-211.