Isolation of a Novel Anti-KDR3 Single-chain Variable Fragment Antibody from a Phage Display Library
-
Shirafkan Kordi
,
Mohammad Rahmati-Yamchi
,
Mehdi Asghari Vostakolaei
,
Ali Etemadie
,
Abolfazl Barzegari
,
Jalal Abdolalizadeh
Abstract
Vascular endothelial growth factor receptor 2 (VEGFR-2) is known as one of the important antigens playing a vital role in angiogenesis. In this study, phage display technology (PDT) was used to produce a single-chain variable fragment (scFv) antibody against a region of the domain 3 in VEGFR-2 called kinase insert domain receptor 3 (KDR3). After designing the KDR3 peptide and biopanning, a colony was chosen for scFv antibody expression. Following expression and purification; western blotting, dot blotting and immunofluorescence (IF) were used to evaluate the antibody function. Surface plasmon resonance (SPR) was also employed to measure affinity of produced antibody. Once a colony was selected and transferred to the expression host, the scFv antibody was expressed in the expected range of 28 kDa. Using a designed chromatography column, antibody purification was found to be about 95%. In this study, a novel scFv with the capability of binding to KDR3 was isolated and purified and its intracellular function was investigated and verified.
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27. Fathi F, Rezabakhsh A, Rahbarghazi R, Rashidi M-R. Early-stage detection of VE-cadherin during endothelial differentiation of human mesenchymal stem cells using SPR biosensor. Biosens Bioelectron 2017; 96:358-66.
28. Fasihi-Ramandi M, Amani J, Salmanian A, Moazzeni S, Ahmadi K. Production and characterization of new anti-human CD20 monoclonal antibody. Iran J Allergy Asthma Immunol 2015; 14(5):502-8.
29. Kordi S, Zarghami N, Akbarzadeh A, Rahmati YM, Ghasemali S, Barkhordari A, et al. A comparison of the inhibitory effect of nano-encapsulated helenalin and free helenalin on telomerase gene expression in the breast cancer cell line, by real-time PCR. Artif Cells Nanomed Biotechnol 2016; 44(2):695-703.
30. Mousavi SA, Ghotaslou R, Kordi S, Khoramdel A, Aeenfar A, Kahjough ST, et al. Antibacterial and antifungal effects of chitosan nanoparticles on tissue conditioners of complete dentures. Int J Biol Macromol 2018; 118:881-5.
31. Akbari B, Farajnia S, Zarghami N, Mahdieh N, Rahmati M, Khosroshahi SA, et al. Design, expression and evaluation of a novel humanized single chain antibody against epidermal growth factor receptor (EGFR). Protein Expr Purif 2016; 127:8-15.
32. Xi H, Yuan R, Chen X, Gu T, Cheng Y, Li Z, et al. Purification and on-column refolding of a single-chain antibody fragment against rabies virus glycoprotein expressed in Escherichia coli. Protein Expr Purif 2016; 126:26-32.
33. Mesgari-Shadi A, Sarrafzadeh MH. Osmotic conditions could promote scFv antibody production in the Escherichia coli HB2151. BioImpacts 2017; 7(3):199-206.
34. Rahbarnia L, Farajnia S, Babaei H, Majidi J, Dariushnejad H, Hosseini MK. Isolation and characterization of a novel human scFv inhibiting EGFR vIII expressing cancers. Immunol Lett 2016; 180:31-8.
35. Vostakolaei MA, Molavi O, Hejazi MS, Kordi S, Rahmati S, Barzegari A, et al. Isolation and characterization of a novel scFv antibody fragments specific for Hsp70 as a tumor biomarker. Journal of cellular biochemistry. 2019.
36. Aghebati-Maleki L, Younesi V, Baradaran B, Abdolalizadeh J, Motallebnezhad M, Nickho H, et al. Antiproliferative and apoptotic effects of novel anti-ROR1 single-chain antibodies in hematological malignancies. SLAS Discov 2017; 22(4):408-17.
37. Akbari B, Farajnia S, Zarghami N, Mahdieh N, Rahmati M, Khosroshahi SA, et al. Construction, expression, and activity of a novel immunotoxin comprising a humanized antiepidermal growth factor receptor scFv and modified Pseudomonas aeruginosa exotoxin A. Anticancer Drugs 2017; 28(3):263-70.
38. Zhang J, Li H, Wang X, Qi H, Miao X, Zhang T, et al. Phage‐derived fully human antibody scFv fragment directed against human vascular endothelial growth factor receptor 2 blocked its interaction with VEGF. Biotechnol Prog 2012; 28(4):981-9.
2. Suto K, Yamazaki Y, Morita T, Mizuno H. Crystal Structures of Novel Vascular Endothelial Growth Factors (VEGF) from Snake Venoms: insight into selective VEGF binding to kinase insert domain-containing receptor but not to fms-like tyrosine kinase-1. J Biol Chem 2005; 280(3):2126-31.
3. Gupta K, Zhang J. Angiogenesis: a curse or cure? Postgrad Med J 2005; 81(954):236-42.
4. Otrock ZK, Makarem JA, Shamseddine AI. Vascular endothelial growth factor family of ligands and receptors: review. Blood Cells Mol Dis 2007; 38(3):258-68.
5. Shibuya M, Claesson-Welsh L. Signal transduction by VEGF receptors in regulation of angiogenesis and lymphangiogenesis. Exp Cell Res 2006; 312(5):549-60.
6. Zhao Y, Adjei AA. Targeting angiogenesis in cancer therapy: moving beyond vascular endothelial growth factor. Oncologist 2015; 20(6):660-73.
7. Hicklin DJ, Ellis LM. Role of the vascular endothelial growth factor pathway in tumor growth and angiogenesis. J Clin Oncol 2005; 23(5):1011-27.
8. Carmeliet P, Jain RK. Molecular mechanisms and clinical applications of angiogenesis. Nature 2011; 473(7347):298-307.
9. McCafferty J, Griffiths AD, Winter G, Chiswell DJ. Phage antibodies: filamentous phage displaying antibody variable domains. Nature 1990; 348(6301):552.
10. Wong KJ, Baidoo KE, Nayak TK, Garmestani K, Brechbiel MW, Milenic DE. In vitro and in vivo pre-clinical analysis of a F (ab') 2 fragment of panitumumab for molecular imaging and therapy of HER1-positive cancers. EJNMMI Res 2011; 1(1):1.
11. Mukai Y, Okamoto T, Kawamura M, Shibata H, Sugita T, Imai S, et al. Optimization of anti-tumor necrosis factor-α single chain Fv displayed on phages for creation of functional antibodies. Pharmazie 2006; 61(10):889-90.
12. Fuh G, Li B, Crowley C, Cunningham B, Wells JA. Requirements for binding and signaling of the kinase domain receptor for vascular endothelial growth factor. J Biol Chem 1998; 273(18):11197-204.
13. Lu D, Kussie P, Pytowski B, Persaud K, Bohlen P, Witte L, et al. Identification of the residues in the extracellular region of KDR important for interaction with vascular endothelial growth factor and neutralizing anti-KDR antibodies. J Biol Chem 2000; 275(19):14321-30.
14. Kordi S, Rahmati-Yamchi M, Vostakolaei MA, Barzegari A, Abdolalizadeh J. Purification of a Novel Anti-VEGFR2 Single Chain Antibody Fragment and Evaluation of Binding Affinity by Surface Plasmon Resonance. Adv Pharm Bull 2019; 9(1):64-9.
15. Larsen JEP, Lund O, Nielsen M. Improved method for predicting linear B-cell epitopes. Immunome Res 2006; 2(1):2.
16. Abdolalizadeh J, Nouri M, Zolbanin JM, Baradaran B, Barzegari A, Omidi Y. Downstream characterization of anti-TNF-α single chain variable fragment antibodies. Hum Antibodies 2012; 21(1-2):41-8.
17. Abdolalizadeh J, Nouri M, Zolbanin JM, Barzegari A, Baradaran B, Barar J, et al. Targeting cytokines: Production and characterization of anti-TNF-α scfvs by phage display technology. Curr Pharm Des 2013; 19(15):2839-47.
18. Kozakov D, Hall DR, Xia B, Porter KA, Padhorny D, Yueh C, et al. The ClusPro web server for protein–protein docking. Nat Protoc 2017; 12(2):255-78.
19. Heo L, Park H, Seok C. GalaxyRefine: protein structure refinement driven by side-chain repacking. Nucleic Acids Res 2013; 41(W1):W384-W8.
20. Lindahl E, Azuara C, Koehl P, Delarue M. NOMAD-Ref: visualization, deformation and refinement of macromolecular structures based on all-atom normal mode analysis. Nucleic Acids Res 2006; 34(suppl_2):W52-W6.
21. Willard L, Ranjan A, Zhang H, Monzavi H, Boyko RF, Sykes BD, et al. VADAR: a web server for quantitative evaluation of protein structure quality. Nucleic Acids Res 2003; 31(13):3316-9.
22. Wiederstein M, Sippl MJ. ProSA-web: interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 2007; 35(suppl_2):W407-W10.
23. Zhang Y, Skolnick J. TM-align: a protein structure alignment algorithm based on the TM-score. Nucleic Acids Res 2005; 33(7):2302-9.
24. Dominguez C, Boelens R, Bonvin AM. HADDOCK: a protein− protein docking approach based on biochemical or biophysical information. J Am Chem Soc 2003; 125(7):1731-7.
25. Abdolalizadeh J, Nouri M, Zolbanin JM, Baradaran B, Barzegari A, Omidi Y. Downstream characterization of anti-TNF-alpha single chain variable fragment antibodies. Hum Antibodies 2012; 21(1-2):41-8.
26. Sharifi M, Dolatabadi JEN, Fathi F, Rashidi M, Jafari B, Tajalli H, et al. Kinetic and thermodynamic study of bovine serum albumin interaction with rifampicin using surface plasmon resonance and molecular docking methods. J Biomed Opt 2017; 22(3):037002.
27. Fathi F, Rezabakhsh A, Rahbarghazi R, Rashidi M-R. Early-stage detection of VE-cadherin during endothelial differentiation of human mesenchymal stem cells using SPR biosensor. Biosens Bioelectron 2017; 96:358-66.
28. Fasihi-Ramandi M, Amani J, Salmanian A, Moazzeni S, Ahmadi K. Production and characterization of new anti-human CD20 monoclonal antibody. Iran J Allergy Asthma Immunol 2015; 14(5):502-8.
29. Kordi S, Zarghami N, Akbarzadeh A, Rahmati YM, Ghasemali S, Barkhordari A, et al. A comparison of the inhibitory effect of nano-encapsulated helenalin and free helenalin on telomerase gene expression in the breast cancer cell line, by real-time PCR. Artif Cells Nanomed Biotechnol 2016; 44(2):695-703.
30. Mousavi SA, Ghotaslou R, Kordi S, Khoramdel A, Aeenfar A, Kahjough ST, et al. Antibacterial and antifungal effects of chitosan nanoparticles on tissue conditioners of complete dentures. Int J Biol Macromol 2018; 118:881-5.
31. Akbari B, Farajnia S, Zarghami N, Mahdieh N, Rahmati M, Khosroshahi SA, et al. Design, expression and evaluation of a novel humanized single chain antibody against epidermal growth factor receptor (EGFR). Protein Expr Purif 2016; 127:8-15.
32. Xi H, Yuan R, Chen X, Gu T, Cheng Y, Li Z, et al. Purification and on-column refolding of a single-chain antibody fragment against rabies virus glycoprotein expressed in Escherichia coli. Protein Expr Purif 2016; 126:26-32.
33. Mesgari-Shadi A, Sarrafzadeh MH. Osmotic conditions could promote scFv antibody production in the Escherichia coli HB2151. BioImpacts 2017; 7(3):199-206.
34. Rahbarnia L, Farajnia S, Babaei H, Majidi J, Dariushnejad H, Hosseini MK. Isolation and characterization of a novel human scFv inhibiting EGFR vIII expressing cancers. Immunol Lett 2016; 180:31-8.
35. Vostakolaei MA, Molavi O, Hejazi MS, Kordi S, Rahmati S, Barzegari A, et al. Isolation and characterization of a novel scFv antibody fragments specific for Hsp70 as a tumor biomarker. Journal of cellular biochemistry. 2019.
36. Aghebati-Maleki L, Younesi V, Baradaran B, Abdolalizadeh J, Motallebnezhad M, Nickho H, et al. Antiproliferative and apoptotic effects of novel anti-ROR1 single-chain antibodies in hematological malignancies. SLAS Discov 2017; 22(4):408-17.
37. Akbari B, Farajnia S, Zarghami N, Mahdieh N, Rahmati M, Khosroshahi SA, et al. Construction, expression, and activity of a novel immunotoxin comprising a humanized antiepidermal growth factor receptor scFv and modified Pseudomonas aeruginosa exotoxin A. Anticancer Drugs 2017; 28(3):263-70.
38. Zhang J, Li H, Wang X, Qi H, Miao X, Zhang T, et al. Phage‐derived fully human antibody scFv fragment directed against human vascular endothelial growth factor receptor 2 blocked its interaction with VEGF. Biotechnol Prog 2012; 28(4):981-9.
| Files | ||
| Issue | Vol 18, No 3 (2019) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijaai.v18i3.1122 | |
| PMID | 31522436 | |
| Keywords | ||
| Monoclonal antibody Kinase insert domain receptor 3 (KDR3) Phage display Single-chain variable fragment (SCFV) Vascular endothelial growth factor receptor-2 (VEGFR2) | ||
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This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
How to Cite
1.
Kordi S, Rahmati-Yamchi M, Asghari Vostakolaei M, Etemadie A, Barzegari A, Abdolalizadeh J. Isolation of a Novel Anti-KDR3 Single-chain Variable Fragment Antibody from a Phage Display Library. Iran J Allergy Asthma Immunol. 2019;18(3):289-299.
