Abstract
Camelid single-domain antibodies (sdAbs, VHHs, or Nanobodies®) are types of antibody fragments that are composed of the heavy-chain variable domain only. These VHHs possess unique structural and functional features, as they are small in size and exhibit thermal stability and high solubility. Compared to conventional antibodies, VHHs can be manufactured in microorganisms to significantly save on cost, labor, and time since VHHs lack the Fc domain with its N-linked oligosaccharide. Until now, VHHs have been expressed in several kinds of production systems, ranging from prokaryotic cells, yeasts, fungi, insect cells, and mammalian cell lines, to plants. In this review, we focus on the recent production of VHHs, introduce different platforms, and summarize the current state of this area and its future trends. Finally, the first potential VHH product, produced in Pichia pastoris, will probably be available on the market in 2018; thus, it is of great importance to give this antibody fragment timely attention. This is the first review concerning the production of VHHs in laboratory settings.
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References
Abe M, Yuki Y, Kurokawa S, Mejima M, Kuroda M, Park EJ, Scheller J, Nakanishi U, Kiyono H (2014) A rice-based soluble form of a murine TNF-specific llama variable domain of heavy-chain antibody suppresses collagen-induced arthritis in mice. J Biotechnol 175:45–52. https://doi.org/10.1016/j.jbiotec.2014.02.005
Agrawal V, Slivac I, Perret S, Bisson L, St-Laurent G, Murad Y, Zhang J, Durocher Y (2012) Stable expression of chimeric heavy chain antibodies in CHO cells. Methods Mol Biol 911:287–303. https://doi.org/10.1007/978-1-61779-968-6_18
Alvarez B, Krogh-Andersen K, Tellgren-Roth C, Martinez N, Gunaydin G, Lin Y, Martin MC, Alvarez MA, Hammarstrom L, Marcotte H (2015) An exopolysaccharide-deficient mutant of Lactobacillus rhamnosus GG efficiently displays a protective llama antibody fragment against rotavirus on its surface. Appl Environ Microbiol 81(17):5784–5793. https://doi.org/10.1128/AEM.00945-15
Alvarez-Rueda N, Behar G, Ferre V, Pugniere M, Roquet F, Gastinel L, Jacquot C, Aubry J, Baty D, Barbet J, Birkle S (2007) Generation of llama single-domain antibodies against methotrexate, a prototypical hapten. Mol Immunol 44(7):1680–1690. https://doi.org/10.1016/j.molimm.2006.08.007
Andersen KK, Strokappe NM, Hultberg A, Truusalu K, Smidt I, Mikelsaar RH, Mikelsaar M, Verrips T, Hammarstrom L, Marcotte H (2015) Neutralization of Clostridium difficile toxin B mediated by engineered Lactobacilli that produce single-domain antibodies. Infect Immun 84(2):395–406. https://doi.org/10.1128/IAI.00870-15
Arbabi Ghahroudi M, Desmyter A, Wyns L, Hamers R, Muyldermans S (1997) Selection and identification of single domain antibody fragments from camel heavy-chain antibodies. FEBS Lett 414(3):521–526. https://doi.org/10.1016/s0014-5793(97)01062-4
Åslund F, Berndt KD, Holmgren A (1997) Redox potentials of glutaredoxins and other thiol-disulfide oxidoreductases of the thioredoxin superfamily determined by direct protein-protein redox equilibria. J Biol Chem 272(49):30780–30786. https://doi.org/10.1074/jbc.272.49.30780
Baghban R, Gargari SL, Rajabibazl M, Nazarian S, Bakherad H (2016) Camelid-derived heavy-chain nanobody against Clostridium botulinum neurotoxin E in Pichia pastoris. Biotechnol Appl Biochem 63(2):200–205. https://doi.org/10.1002/bab.1226
Bakherad H, Mousavi Gargari SL, Rasooli I, Rajabibazl M, Mohammadi M, Ebrahimizadeh W, Safaee Ardakani L, Zare H (2013) In vivo neutralization of botulinum neurotoxins serotype E with heavy-chain camelid antibodies (VHH). Mol Biotechnol 55(2):159–167. https://doi.org/10.1007/s12033-013-9669-1
Bazl MR, Rasaee MJ, Foruzandeh M, Rahimpour A, Kiani J, Rahbarizadeh F, Alirezapour B, Mohammadi M (2007) Production of chimeric recombinant single domain antibody-green fluorescent fusion protein in Chinese hamster ovary cells. Hybridoma 26(1):1–9. https://doi.org/10.1089/hyb.2006.037
Behdani M, Zeinali S, Karimipour M, Khanahmad H, Schoonooghe S, Aslemarz A, Seyed N, Moazami-Godarzi R, Baniahmad F, Habibi-Anbouhi M, Hassanzadeh-Ghassabeh G, Muyldermans S (2013) Development of VEGFR2-specific nanobody pseudomonas exotoxin A conjugated to provide efficient inhibition of tumor cell growth. New Biotechnol 30(2):205–209. https://doi.org/10.1016/j.nbt.2012.09.002
Bessette PH, Åslund F, Beckwith J, Georgiou G (1999) Efficient folding of proteins with multiple disulfide bonds in the Escherichia coli cytoplasm. Proc Natl Acad Sci U S A 96(24):13703–13708. https://doi.org/10.1073/pnas.96.24.13703
Billen B, Vincke C, Hansen R, Devoogdt N, Muyldermans S, Adriaensens P, Guedens W (2017) Cytoplasmic versus periplasmic expression of site-specifically and bioorthogonally functionalized nanobodies using expressed protein ligation. Protein Expr Purif 133:25–34. https://doi.org/10.1016/j.pep.2017.02.009
Bossi S, Ferranti B, Martinelli C, Capasso P, de Marco A (2010) Antibody-mediated purification of co-expressed antigen–antibody complexes. Protein Expr Purif 72(1):55–58. https://doi.org/10.1016/j.pep.2010.01.003
Conrad U, Plagmann I, Malchow S, Sack M, Floss DM, Kruglov AA, Nedospasov SA, Rose-John S, Scheller J (2011) ELPylated anti-human TNF therapeutic single-domain antibodies for prevention of lethal septic shock. Plant Biotechnol J 9(1):22–31. https://doi.org/10.1111/j.1467-7652.2010.00523.x
De Buck S, Nolf J, De Meyer T, Virdi V, De Wilde K, Van Lerberge E, Van Droogenbroeck B, Depicker A (2013) Fusion of an Fc chain to a VHH boosts the accumulation levels in Arabidopsis seeds. Plant Biotechnol J 11(8):1006–1016. https://doi.org/10.1111/pbi.12094
De Meyer T, Laukens B, Nolf J, Van Lerberge E, De Rycke R, De Beuckelaer A, De Buck S, Callewaert N, Depicker A (2015) Comparison of VHH-Fc antibody production in Arabidopsis thaliana, Nicotiana benthamiana and Pichia pastoris. Plant Biotechnol J 13(7):938–947. https://doi.org/10.1111/pbi.12330
Djender S, Schneider A, Beugnet A, Crepin R, Desrumeaux KE, Romani C, Moutel S, Perez F, de Marco A (2014) Bacterial cytoplasm as an effective cell compartment for producing functional VHH-based affinity reagents and Camelidae IgG-like recombinant antibodies. Microb Cell Factories 13(1):140. https://doi.org/10.1186/s12934-014-0140-1
Dolk E, van Vliet C, Perez JM, Vriend G, Darbon H, Ferrat G, Cambillau C, Frenken LG, Verrips T (2005) Induced refolding of a temperature denatured llama heavy-chain antibody fragment by its antigen. Proteins 59(3):555–564. https://doi.org/10.1002/prot.20378
Dumoulin M, Last AM, Desmyter A, Decanniere K, Canet D, Larsson G, Spencer A, Archer DB, Sasse J, Muyldermans S, Wyns L, Redfield C, Matagne A, Robinson CV, Dobson CM (2003) A camelid antibody fragment inhibits the formation of amyloid fibrils by human lysozyme. Nature 424(6950):783–788. https://doi.org/10.1038/nature01870
Ezzine A, M’ Hirsi El Adab S, Bouhaouala-Zahar B, Hmila I, Baciou L, Marzouki MN (2012) Efficient expression of the anti-AahI’ scorpion toxin nanobody under a new functional form in a Pichia pastoris system. Biotechnol Appl Biochem 59(1):15–21. https://doi.org/10.1002/bab.67
Farasat A, Rahbarizadeh F, Ahmadvand D, Yazdian F (2017) Optimization of an anti-HER2 nanobody expression using the Taguchi method. Prep Biochem Biotechnol 47(8):795–803. https://doi.org/10.1080/10826068.2017.1342259
Frenken LGJ, van der Linden RHJ, Hermans PWJJ, Bos JW, Ruuls RC, de Geus B, Verrips CT (2000) Isolation of antigen specific llama VHH antibody fragments and their high level secretion by Saccharomyces cerevisiae. J Biotechnol 78(1):11–21. https://doi.org/10.1016/S0168-1656(99)00228-X
Giuliani M, Parrilli E, Sannino F, Apuzzo G, Marino G, Tutino ML (2015) Soluble recombinant protein production in Pseudoalteromonas haloplanktis TAC125. In: García-Fruitós E (ed) Insoluble proteins: methods and protocols. Springer New York, New York, pp 243–257
Goldman ER, Brozozog-Lee PA, Zabetakis D, Turner KB, Walper SA, Liu JL, Anderson GP (2014) Negative tail fusions can improve ruggedness of single domain antibodies. Protein Expr Purif 95:226–232. https://doi.org/10.1016/j.pep.2014.01.003
Gómez-Sebastián S, Nuñez MC, Garaicoechea L, Alvarado C, Mozgovoj M, Lasa R, Kahl A, Wigdorovitz A, Parreño V, Escribano JM (2012) Rotavirus A-specific single-domain antibodies produced in baculovirus-infected insect larvae are protective in vivo. BMC Biotechnol 12(1):59–59. https://doi.org/10.1186/1472-6750-12-59
Gorlani A, de Haard H, Verrips T (2012a) Expression of VHHs in Saccharomyces cerevisiae. Methods Mol Biol 911:277–286. https://doi.org/10.1007/978-1-61779-968-6_17
Gorlani A, Hulsik DL, Adams H, Vriend G, Hermans P, Verrips T (2012b) Antibody engineering reveals the important role of J segments in the production efficiency of llama single-domain antibodies in Saccharomyces cerevisiae. Protein Eng Des Sel: PEDS 25(1):39–46. https://doi.org/10.1093/protein/gzr057
Günaydın G, Álvarez B, Lin Y, Hammarström L, Marcotte H (2014) Co-expression of anti-rotavirus proteins (llama VHH antibody fragments) in Lactobacillus: development and functionality of vectors containing two expression cassettes in tandem. PLoS One 9(4):e96409. https://doi.org/10.1371/journal.pone.0096409
Hamers-Casterman C, Atarhouch T, Muyldermans S, Robinson G, Hamers C, Songa EB, Bendahman N, Hamers R (1993) Naturally occurring antibodies devoid of light chains. Nature 363(6428):446–448. https://doi.org/10.1038/363446a0
Harmsen MM, Ruuls RC, Nijman IJ, Niewold TA, Frenken LGJ, de Geus B (2000) Llama heavy-chain V regions consist of at least four distinct subfamilies revealing novel sequence features. Mol Immunol 37(10):579–590. https://doi.org/10.1016/S0161-5890(00)00081-X
Harmsen MM, Van Solt CB, Fijten HP, Van Setten MC (2005) Prolonged in vivo residence times of llama single-domain antibody fragments in pigs by binding to porcine immunoglobulins. Vaccine 23(41):4926–4934. https://doi.org/10.1016/j.vaccine.2005.05.017
Harmsen MM, van Solt CB, Fijten HP (2009) Enhancement of toxin- and virus-neutralizing capacity of single-domain antibody fragments by N-glycosylation. Appl Microbiol Biotechnol 84(6):1087–1094. https://doi.org/10.1007/s00253-009-2029-1
Hatahet F, Nguyen VD, Salo KE, Ruddock LW (2010) Disruption of reducing pathways is not essential for efficient disulfide bond formation in the cytoplasm of E. coli. Microb Cell Factories 9:67. https://doi.org/10.1186/1475-2859-9-67
Hellens RP, Allan AC, Friel EN, Bolitho K, Grafton K, Templeton MD, Karunairetnam S, Gleave AP, Laing WA (2005) Transient expression vectors for functional genomics, quantification of promoter activity and RNA silencing in plants. Plant Methods 1(1):13. https://doi.org/10.1186/1746-4811-1-13
Henry KA, Sulea T, van Faassen H, Hussack G, Purisima EO, MacKenzie CR, Arbabi-Ghahroudi M (2016) A rational engineering strategy for designing protein A-binding camelid single-domain antibodies. PLoS One 11(9):e0163113. https://doi.org/10.1371/journal.pone.0163113
Hirayama K, Watanabe H, Tokuda G, Kitamoto K, Arioka M (2010) Purification and characterization of termite endogenous beta-1,4-endoglucanases produced in Aspergillus oryzae. Biosci Biotechnol Biochem 74(8):1680–1686. https://doi.org/10.1271/bbb.100296
Hisada H, Tsutsumi H, Ishida H, Hata Y (2013) High production of llama variable heavy-chain antibody fragment (VHH) fused to various reader proteins by Aspergillus oryzae. Appl Microbiol Biotechnol 97(2):761–766. https://doi.org/10.1007/s00253-012-4211-0
Hmila I, Abdallah RB, Saerens D, Benlasfar Z, Conrath K, Ayeb ME, Muyldermans S, Bouhaouala-Zahar B (2008) VHH, bivalent domains and chimeric heavy chain-only antibodies with high neutralizing efficacy for scorpion toxin AahI’. Mol Immunol 45(14):3847–3856. https://doi.org/10.1016/j.molimm.2008.04.011
Hussack G, Arbabi-Ghahroudi M, van Faassen H, Songer JG, Ng KK, MacKenzie R, Tanha J (2011) Neutralization of Clostridium difficile toxin A with single-domain antibodies targeting the cell receptor binding domain. J Biol Chem 286(11):8961–8976. https://doi.org/10.1074/jbc.M110.198754
Ismaili A, Jalali-Javaran M, Rasaee Mohammad J, Rahbarizadeh F, Forouzandeh-Moghadam M, Memari Hamid R (2007) Production and characterization of anti-(mucin MUC1) single-domain antibody in tobacco (Nicotiana tabacum cultivar Xanthi). Biotechnol Appl Biochem 47(1):11–19. https://doi.org/10.1042/BA20060071
Jarviluoma A, Strandin T, Lulf S, Bouchet J, Makela AR, Geyer M, Benichou S, Saksela K (2012) High-affinity target binding engineered via fusion of a single-domain antibody fragment with a ligand-tailored SH3 domain. PLoS One 7(7):e40331. https://doi.org/10.1371/journal.pone.0040331
Ji X, Lu W, Zhou H, Han D, Yang L, Wu H, Li J, Liu H, Zhang J, Cao P, Zhang S (2013) Covalently dimerized Camelidae antihuman TNFa single-domain antibodies expressed in yeast Pichia pastoris show superior neutralizing activity. Appl Microbiol Biotechnol 97(19):8547–8558. https://doi.org/10.1007/s00253-012-4639-2
Jobling SA, Jarman C, Teh MM, Holmberg N, Blake C, Verhoeyen ME (2003) Immunomodulation of enzyme function in plants by single-domain antibody fragments. Nat Biotechnol 21(1):77–80. https://doi.org/10.1038/nbt772
Joosten V, Gouka RJ, van den Hondel CA, Verrips CT, Lokman BC (2005a) Expression and production of llama variable heavy-chain antibody fragments (V(HH)s) by Aspergillus awamori. Appl Microbiol Biotechnol 66(4):384–392. https://doi.org/10.1007/s00253-004-1689-0
Joosten V, Roelofs MS, van den Dries N, Goosen T, Verrips CT, van den Hondel CA, Lokman BC (2005b) Production of bifunctional proteins by Aspergillus awamori: llama variable heavy chain antibody fragment (V(HH)) R9 coupled to Arthromyces ramosus peroxidase (ARP). J Biotechnol 120(4):347–359. https://doi.org/10.1016/j.jbiotec.2005.06.034
Koch-Nolte F, Reyelt J, Schossow B, Schwarz N, Scheuplein F, Rothenburg S, Haag F, Alzogaray V, Cauerhff A, Goldbaum FA (2007) Single domain antibodies from llama effectively and specifically block T cell ecto-ADP-ribosyltransferase ART2.2 in vivo. FASEB J: Ofl Publ Fed Am Soc Exp Biol 21(13):3490–3498. https://doi.org/10.1096/fj.07-8661com
Lentz EM, Garaicoechea L, Alfano EF, Parreno V, Wigdorovitz A, Bravo-Almonacid FF (2012) Translational fusion and redirection to thylakoid lumen as strategies to improve the accumulation of a camelid antibody fragment in transplastomic tobacco. Planta 236(2):703–714. https://doi.org/10.1007/s00425-012-1642-x
Li M, Fan X, Liu J, Hu Y, Huang H (2015) Selection by phage display of nanobodies directed against hypoxia inducible factor-1alpha (HIF-1alpha). Biotechnol Appl Biochem 62(6):738–745. https://doi.org/10.1002/bab.1340
Maggi M, Scotti C (2017) Enhanced expression and purification of camelid single domain VHH antibodies from classical inclusion bodies. Protein Expr Purif 136:39–44. https://doi.org/10.1016/j.pep.2017.02.007
Makvandi-Nejad S, Fjallman T, Arbabi-Ghahroudi M, MacKenzie CR, Hall JC (2011) Selection and expression of recombinant single domain antibodies from a hyper-immunized library against the hapten azoxystrobin. J Immunol Methods 373(1–2):8–18. https://doi.org/10.1016/j.jim.2011.07.006
Marcobal A, Liu X, Zhang W, Dimitrov AS, Jia L, Lee PP, Fouts TR, Parks TP, Lagenaur LA (2016) Expression of human immunodeficiency virus type 1 neutralizing antibody fragments using human vaginal Lactobacillus. AIDS Res Hum Retrovir 32(10–11):964–971. https://doi.org/10.1089/AID.2015.0378
Markiv A, Beatson R, Burchell J, Durvasula RV, Kang AS (2011) Expression of recombinant multi-coloured fluorescent antibodies in gor -/trxB- E. coli cytoplasm. BMC Biotechnol 11(1):117. https://doi.org/10.1186/1472-6750-11-117
Martin MC, Pant N, Ladero V, Gunaydin G, Andersen KK, Alvarez B, Martinez N, Alvarez MA, Hammarstrom L, Marcotte H (2011) Integrative expression system for delivery of antibody fragments by lactobacilli. Appl Environ Microbiol 77(6):2174–2179. https://doi.org/10.1128/AEM.02690-10
Maskos K, Huber-Wunderlich M, Glockshuber R (2003) DsbA and DsbC-catalyzed oxidative folding of proteins with complex disulfide bridge patterns in vitro and in vivo. J Mol Biol 325(3):495–513. https://doi.org/10.1016/s0022-2836(02)01248-2
McCue JT, Selvitelli K, Walker J (2009) Application of a novel affinity adsorbent for the capture and purification of recombinant factor VIII compounds. J Chromatogr A 1216(45):7824–7830. https://doi.org/10.1016/j.chroma.2009.09.045
McLaughlin P, Grillo-López AJ, Link BK, Levy R, Czuczman MS, Williams ME, Heyman MR, Bence-Bruckler I, White CA, Cabanillas F, Jain V, Ho AD, Lister J, Wey K, Shen D, Dallaire BK (1998) Rituximab chimeric anti-CD20 monoclonal antibody therapy for relapsed indolent lymphoma: half of patients respond to a four-dose treatment program. J Clin Oncol 16(8):2825–2833. https://doi.org/10.1200/jco.1998.16.8.2825
Missiakas D, Betton J-M, Raina S (1996) New components of protein folding in extracytoplasmic compartments of Escherichia coli SurA, FkpA and Skp/OmpH. Mol Microbiol 21(4):871–884. https://doi.org/10.1046/j.1365-2958.1996.561412.x
Mizukami M, Tokunaga H, Onishi H, Ueno Y, Hanagata H, Miyazaki N, Kiyose N, Ito Y, Ishibashi M, Hagihara Y, Arakawa T, Miyauchi A, Tokunaga M (2015) Highly efficient production of VHH antibody fragments in Brevibacillus choshinensis expression system. Protein Expr Purif 105:23–32. https://doi.org/10.1016/j.pep.2014.09.017
Nettleship JE, Ren J, Rahman N, Berrow NS, Hatherley D, Barclay AN, Owens RJ (2008) A pipeline for the production of antibody fragments for structural studies using transient expression in HEK 293T cells. Protein Expr Purif 62(1):83–89. https://doi.org/10.1016/j.pep.2008.06.017
Neu HC, Heppel LA (1965) The release of enzymes from Escherichia coli by osmotic shock and during the formation of spheroplasts. J Biol Chem 240(9):3685–3692
Nguyen VK, Zou X, Lauwereys M, Brys L, Brüggemann M, Muyldermans S (2003) Heavy-chain only antibodies derived from dromedary are secreted and displayed by mouse B cells. Immunology 109(1):93–101. https://doi.org/10.1046/j.1365-2567.2003.01633.x
Noguchi T, Nishida Y, Takizawa K, Cui Y, Tsutsumi K, Hamada T, Nishi Y (2017) Accurate quantitation for in vitro refolding of single domain antibody fragments expressed as inclusion bodies by referring the concomitant expression of a soluble form in the periplasms of Escherichia coli. J Immunol Methods 442:1–11. https://doi.org/10.1016/j.jim.2016.11.014
Nozach H, Fruchart-Gaillard C, Fenaille F, Beau F, Ramos OHP, Douzi B, Saez NJ, Moutiez M, Servent D, Gondry M, Thaï R, Cuniasse P, Vincentelli R, Dive V (2013) High throughput screening identifies disulfide isomerase DsbC as a very efficient partner for recombinant expression of small disulfide-rich proteins in E. coli. Microb Cell Factories 12(1):37–37. https://doi.org/10.1186/1475-2859-12-37
Nyyssonen E, Penttila M, Harkki A, Saloheimo A, Knowles JK, Keranen S (1993) Efficient production of antibody fragments by the filamentous fungus Trichoderma reesei. Bio/Technol (Nat Publ Co) 11(5):591–595
Okazaki F, Aoki J, Tabuchi S, Tanaka T, Ogino C, Kondo A (2012) Efficient heterologous expression and secretion in Aspergillus oryzae of a llama variable heavy-chain antibody fragment V(HH) against EGFR. Appl Microbiol Biotechnol 96(1):81–88. https://doi.org/10.1007/s00253-012-4158-1
Olichon A, Surrey T (2007) Selection of genetically encoded fluorescent single domain antibodies engineered for efficient expression in Escherichia coli. J Biol Chem 282(50):36314–36320. https://doi.org/10.1074/jbc.M704908200
Olichon A, Schweizer D, Muyldermans S, de Marco A (2007) Heating as a rapid purification method for recovering correctly-folded thermotolerant VH and VHH domains. BMC Biotechnol 7(1):7. https://doi.org/10.1186/1472-6750-7-7
Omidfar K, Rasaee MJ, Kashanian S, Paknejad M, Bathaie Z (2007) Studies of thermostability in Camelus bactrianus (Bactrian camel) single-domain antibody specific for the mutant epidermal-growth-factor receptor expressed by Pichia. Biotechnol Appl Biochem 46(Pt 1):41–49. https://doi.org/10.1042/BA20060104
Paal M, Heel T, Schneider R, Auer B (2009) A novel Ecotin-Ubiquitin-Tag (ECUT) for efficient, soluble peptide production in the periplasm of Escherichia coli. Microb Cell Factories 8(1):7. https://doi.org/10.1186/1475-2859-8-7
Pant N, Hultberg A, Zhao Y, Svensson L, Pan-Hammarström Q, Johansen K, Pouwels PH, Ruggeri FM, Hermans P, Frenken L, Borén T, Marcotte H, Hammarström L (2006) Lactobacilli expressing variable domain of llama heavy-chain antibody fragments (lactobodies) confer protection against rotavirus-induced diarrhea. J Infect Dis 194(11):1580–1588. https://doi.org/10.1086/508747
Pant N, Marcotte H, Hermans P, Bezemer S, Frenken L, Johansen K, Hammarström L (2011) Lactobacilli producing bispecific llama-derived anti-rotavirus proteins in vivo for rotavirus-induced diarrhea. Future Microbiol 6(5):583–593. https://doi.org/10.2217/fmb.11.32
Proba K, Ge L, Plückthun A (1995) Functional antibody single-chain fragments from the cytoplasm of Escherichia coli: influence of thioredoxin reductase (TrxB). Gene 159(2):203–207. https://doi.org/10.1016/0378-1119(95)00018-2
Qasemi M, Behdani M, Shokrgozar MA, Molla-Kazemiha V, Mohseni-Kuchesfahani H, Habibi-Anbouhi M (2016) Construction and expression of an anti-VEGFR2 nanobody-Fc fusionbody in NS0 host cell. Protein Expr Purif 123:19–25. https://doi.org/10.1016/j.pep.2016.03.004
Rahbarizadeh F, Rasaee MJ, Forouzandeh-Moghadam M, Allameh AA (2005) High expression and purification of the recombinant camelid anti-MUC1 single domain antibodies in Escherichia coli. Protein Expr Purif 44(1):32–38. https://doi.org/10.1016/j.pep.2005.04.008
Rahbarizadeh F, Rasaee MJ, Forouzandeh M, Allameh AA (2006) Over expression of anti-MUC1 single-domain antibody fragments in the yeast Pichia pastoris. Mol Immunol 43(5):426–435. https://doi.org/10.1016/j.molimm.2005.03.003
Rajabi-Memari H, Jalali-Javaran M, Rasaee MJ, Rahbarizadeh F, Forouzandeh-Moghadam M, Esmaili A (2006) Expression and characterization of a recombinant single-domain monoclonal antibody against MUC1 mucin in tobacco plants. Hybridoma 25(4):209–215. https://doi.org/10.1089/hyb.2006.25.209
Richard G, Meyers AJ, McLean MD, Arbabi-Ghahroudi M, MacKenzie R, Hall JC (2013) In vivo neutralization of alpha-cobratoxin with high-affinity llama single-domain antibodies (VHHs) and a VHH-Fc antibody. PLoS One 8(7):e69495. https://doi.org/10.1371/journal.pone.0069495
Rietsch A, Belin D, Martin N, Beckwith J (1996) An in vivo pathway for disulfide bond isomerization in Escherichia coli. Proc Natl Acad Sci U S A 93(23):13048–13053. https://doi.org/10.1073/pnas.93.23.13048
Roovers RC, Vosjan MJ, Laeremans T, el Khoulati R, de Bruin RC, Ferguson KM, Verkleij AJ, van Dongen GA, van Bergen en Henegouwen PM (2011) A biparatopic anti-EGFR nanobody efficiently inhibits solid tumour growth. Int J Cancer 129(8):2013–2024. https://doi.org/10.1002/ijc.26145
Rotman M, Welling MM, van den Boogaard ML, Moursel LG, van der Graaf LM, van Buchem MA, van der Maarel SM, van der Weerd L (2015) Fusion of hIgG1-Fc to In-111-anti-amyloid single domain antibody fragment VHH-pa2H prolongs blood residential time in APP/PS1 mice but does not increase brain uptake. Nucl Med Biol 42(8):695–702. https://doi.org/10.1016/j.nucmedbio.2015.03.003
Sagt CMJ, Kleizen B, Verwaal R, de Jong MDM, Müller WH, Smits A, Visser C, Boonstra J, Verkleij AJ, Verrips CT (2000) Introduction of an N-glycosylation site increases secretion of heterologous proteins in yeasts. Appl Environ Microbiol 66(11):4940–4944. https://doi.org/10.1128/aem.66.11.4940-4944.2000
Schlegel S, Rujas E, Ytterberg AJ, Zubarev RA, Luirink J, de Gier J-W (2013) Optimizing heterologous protein production in the periplasm of E. coli by regulating gene expression levels. Microb Cell Factories 12(1):24–24. https://doi.org/10.1186/1475-2859-12-24
Shkoporov AN, Khokhlova EV, Savochkin KA, Kafarskaia LI, Efimov BA (2015) Production of biologically active scFv and VHH antibody fragments in Bifidobacterium longum. FEMS Microbiol Lett 362(12):fnv083. https://doi.org/10.1093/femsle/fnv083
Shriver-Lake LC, Goldman ER, Zabetakis D, Anderson GP (2017) Improved production of single domain antibodies with two disulfide bonds by co-expression of chaperone proteins in the Escherichia coli periplasm. J Immunol Methods 443:64–67. https://doi.org/10.1016/j.jim.2017.01.007
Sletta H, Tondervik A, Hakvag S, Aune TE, Nedal A, Aune R, Evensen G, Valla S, Ellingsen TE, Brautaset T (2007) The presence of N-terminal secretion signal sequences leads to strong stimulation of the total expression levels of three tested medically important proteins during high-cell-density cultivations of Escherichia coli. Appl Environ Microbiol 73(3):906–912. https://doi.org/10.1128/aem.01804-06
Strasser R, Altmann F, Steinkellner H (2014) Controlled glycosylation of plant-produced recombinant proteins. Curr Opin Biotechnol 30:95–100. https://doi.org/10.1016/j.copbio.2014.06.008
Ta DT, Redeker ES, Billen B, Reekmans G, Sikulu J, Noben JP, Guedens W, Adriaensens P (2015) An efficient protocol towards site-specifically clickable nanobodies in high yield: cytoplasmic expression in Escherichia coli combined with intein-mediated protein ligation. Protein Eng, Des Sel: PEDS 28(10):351–363. https://doi.org/10.1093/protein/gzv032
Teh YH, Kavanagh TA (2010) High-level expression of camelid nanobodies in Nicotiana benthamiana. Transgenic Res 19(4):575–586. https://doi.org/10.1007/s11248-009-9338-0
Terfruchte M, Reindl M, Jankowski S, Sarkari P, Feldbrugge M, Schipper K (2017) Applying unconventional secretion in Ustilago maydis for the export of functional nanobodies. Int J Mol Sci 18(5):937. https://doi.org/10.3390/ijms18050937
Thomassen YE, Meijer W, Sierkstra L, Verrips CT (2002) Large-scale production of VHH antibody fragments by Saccharomyces cerevisiae. Enzym Microb Technol 30(3):273–278. https://doi.org/10.1016/S0141-0229(01)00497-5
Thomassen YE, Verkleij AJ, Boonstra J, Verrips CT (2005) Specific production rate of VHH antibody fragments by Saccharomyces cerevisiae is correlated with growth rate, independent of nutrient limitation. J Biotechnol 118(3):270–277. https://doi.org/10.1016/j.jbiotec.2005.05.010
Uttamchandani M, Neo JL, Ong BN, Moochhala S (2009) Applications of microarrays in pathogen detection and biodefence. Trends Biotechnol 27(1):53–61. https://doi.org/10.1016/j.tibtech.2008.09.004
van den Hombergh JPTW, van de Vondervoort PJI, Fraissinet-Tachet L, Visser J (1997) Aspergillus as a host for heterologous protein production: the problem of proteases. Trends Biotechnol 15(7):256–263. https://doi.org/10.1016/S0167-7799(97)01020-2
van der Linden RHJ, de Geus B, Frenken LGJ, Peters H, Verrips CT (2000) Improved production and function of llama heavy chain antibody fragments by molecular evolution. J Biotechnol 80(3):261–270. https://doi.org/10.1016/S0168-1656(00)00274-1
van der Vaart JM, Pant N, Wolvers D, Bezemer S, Hermans PW, Bellamy K, Sarker SA, van der Logt CPE, Svensson L, Verrips CT, Hammarstrom L, van Klinken BJW (2006) Reduction in morbidity of rotavirus induced diarrhoea in mice by yeast produced monovalent llama-derived antibody fragments. Vaccine 24(19):4130–4137. https://doi.org/10.1016/j.vaccine.2006.02.045
Vandenbroucke K, de Haard H, Beirnaert E, Dreier T, Lauwereys M, Huyck L, Van Huysse J, Demetter P, Steidler L, Remaut E, Cuvelier C, Rottiers P (2010) Orally administered L. lactis secreting an anti-TNF nanobody demonstrate efficacy in chronic colitis. Mucosal Immunol 3(1):49–56. https://doi.org/10.1038/mi.2009.116
Vaneycken I, Devoogdt N, Van Gassen N, Vincke C, Xavier C, Wernery U, Muyldermans S, Lahoutte T, Caveliers V (2011) Preclinical screening of anti-HER2 nanobodies for molecular imaging of breast cancer. FASEB J: Off Publ Fed Am Soc Exp Biol 25(7):2433–2446. https://doi.org/10.1096/fj.10-180331
Vee Aune TE, Bakke I, Drablos F, Lale R, Brautaset T, Valla S (2010) Directed evolution of the transcription factor XylS for development of improved expression systems. Microb Biotechnol 3(1):38–47. https://doi.org/10.1111/j.1751-7915.2009.00126.x
Veggiani G, de Marco A (2011) Improved quantitative and qualitative production of single-domain intrabodies mediated by the co-expression of Erv1p sulfhydryl oxidase. Protein Expr Purif 79(1):111–114. https://doi.org/10.1016/j.pep.2011.03.005
Virdi V, Coddens A, De Buck S, Millet S, Goddeeris BM, Cox E, De Greve H, Depicker A (2013) Orally fed seeds producing designer IgAs protect weaned piglets against enterotoxigenic Escherichia coli infection. Proc Natl Acad Sci U S A 110(29):11809–11814. https://doi.org/10.1073/pnas.1301975110
Wagner R, Liedtke S, Kretzschmar E, Geyer H, Geyer R, Klenk H-D (1996) Elongation of the N-glycans of fowl plague virus hemagglutinin expressed in Spodoptera frugiperda (Sf9) cells by coexpression of human β1,2-N-acetylglucosaminyltransferase I. Glycobiology 6(2):165–175. https://doi.org/10.1093/glycob/6.2.165
Wesolowski J, Alzogaray V, Reyelt J, Unger M, Juarez K, Urrutia M, Cauerhff A, Danquah W, Rissiek B, Scheuplein F, Schwarz N, Adriouch S, Boyer O, Seman M, Licea A, Serreze DV, Goldbaum FA, Haag F, Koch-Nolte F (2009) Single domain antibodies: promising experimental and therapeutic tools in infection and immunity. Med Microbiol Immunol 198(3):157–174. https://doi.org/10.1007/s00430-009-0116-7
Winichayakul S, Pernthaner A, Scott R, Vlaming R, Roberts N (2009) Head-to-tail fusions of camelid antibodies can be expressed in planta and bind in rumen fluid. Biotechnol Appl Biochem 53(2):111–122. https://doi.org/10.1042/BA20080076
Xue X, Fan X, Qu Q, Wu G (2016) Bioscreening and expression of a camel anti-CTGF VHH nanobody and its renaturation by a novel dialysis-dilution method. AMB Express 6(1):72. https://doi.org/10.1186/s13568-016-0249-1
Yang Z, Schmidt D, Liu W, Li S, Shi L, Sheng J, Chen K, Yu H, Tremblay JM, Chen X, Piepenbrink KH, Sundberg EJ, Kelly CP, Bai G, Shoemaker CB, Feng H (2014) A novel multivalent, single-domain antibody targeting TcdA and TcdB prevents fulminant Clostridium difficile infection in mice. J Infect Dis 210(6):964–972. https://doi.org/10.1093/infdis/jiu196
Zarschler K, Witecy S, Kapplusch F, Foerster C, Stephan H (2013) High-yield production of functional soluble single-domain antibodies in the cytoplasm of Escherichia coli. Microb Cell Factories 12(1):97–97. https://doi.org/10.1186/1475-2859-12-97
Zasada AA, Rastawicki W, Śmietańska K, Rokosz N, Jagielski M (2013) Comparison of seven commercial enzyme-linked immunosorbent assays for the detection of anti-diphtheria toxin antibodies. Eur J Clin Microbiol Infect Dis 32(7):891–897. https://doi.org/10.1007/s10096-013-1823-y
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This work was supported by the National Natural Science Foundation of China (Grant No. 31470967).
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Liu, Y., Huang, H. Expression of single-domain antibody in different systems. Appl Microbiol Biotechnol 102, 539–551 (2018). https://doi.org/10.1007/s00253-017-8644-3
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DOI: https://doi.org/10.1007/s00253-017-8644-3