Improved Fmoc-solid-phase peptide synthesis of an extracellular loop of CFTR for antibody selection by the phage display technology.
CFTR
PEGylation
aspartimide
biotinylation
cyclization
cystic fibrosis
extracellular loop
solid-phase peptide synthesis
Journal
Journal of peptide science : an official publication of the European Peptide Society
ISSN: 1099-1387
Titre abrégé: J Pept Sci
Pays: England
ID NLM: 9506309
Informations de publication
Date de publication:
Jul 2020
Jul 2020
Historique:
received:
20
03
2020
revised:
20
04
2020
accepted:
24
04
2020
pubmed:
14
5
2020
medline:
9
2
2021
entrez:
14
5
2020
Statut:
ppublish
Résumé
Cystic fibrosis (CF), a life-shortening genetic disease, is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene that codes for the CFTR protein, the major chloride channel expressed at the apical membrane of epithelial cells. The development of an imaging probe capable of non-invasively detect CFTR at the cell surface could be of great advantage for the management of CF. With that purpose, we synthesized the first extracellular loop of CFTR protein (ECL1) through fluorenylmethyloxycarbonyl (Fmoc)-based microwave-assisted solid-phase peptide synthesis (SPPS), according to a reported methodology. However, aspartimide formation, a well-characterized side reaction in Fmoc-SPPS, prompted us to adopt a different side-chain protection strategy for aspartic acid residues present in ECL1 sequence. The peptide was subsequently modified via PEGylation and biotinylation, and cyclized through disulfide bridge formation, mimicking the native loop conformation in CFTR protein. Herein, we report improvements in the synthesis of the first extracellular loop of CFTR, including peptide modifications that can be used to improve antigen presentation in phage display for selection of novel antibodies against plasma membrane CFTR.
Substances chimiques
9-fluorenylmethoxycarbonyl
0
Antibodies
0
CFTR protein, human
0
Fluorenes
0
Peptides
0
Cystic Fibrosis Transmembrane Conductance Regulator
126880-72-6
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e3253Subventions
Organisme : BioISI
ID : UID/MULTI/04046/2019
Organisme : C2TN
ID : UID/Multi/04349/2019
Organisme : Lisboa2020-EU FEDER
Organisme : Fundação para a Ciência e Tecnologia (FCT)
ID : PTDC/BTM-TEC/29256/2017
Organisme : Fundação para a Ciência e Tecnologia (FCT)
ID : LISBOA-01-0145-FEDER-022125 from FCT and Lisboa2020-EU FEDER
Informations de copyright
© 2020 European Peptide Society and John Wiley & Sons, Ltd.
Références
Riordan JR, Rommens JM, Kerem B, et al. Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. Science. 1989;245(4922):1066-1073.
Cystic Fibrosis Mutation Database homepage. , http://www.genet.sickkids.on.ca/Home.html (accessed: April 2020)
Van Goor F, Hadida S, Grootenhuis PDJ, et al. Rescue of CF airway epithelial cell function in vitro by a CFTR potentiator, VX-770. Proc Natl Acad Sci U S A. 2009;106(44):18825-18830.
Ramsey BW, Davies J, McElvaney NG, et al. A CFTR potentiator in patients with cystic fibrosis and the G551D mutation. N Engl J Med. 2011;365:1663-1672.
Davies JC, Wainwright CE, Canny GJ, et al. Efficacy and safety of ivacaftor in patients aged 6 to 11 years with cystic fibrosis with a G551D mutation. Am J Respir Crit Care Med. 2013;187(11):1219-1225.
De Boeck K, Munck A, Walker S, et al. Efficacy and safety of ivacaftor in patients with cystic fibrosis and a non-G551D gating mutation. J Cyst Fibros. 2014;13(6):674-680.
McKone EF, Borowitz D, Drevinek P, et al. Long-term safety and efficacy of ivacaftor in patients with cystic fibrosis who have the Gly551Asp-CFTR mutation: a phase 3, open-label extension study (PERSIST). Lancet Respir Med. 2014;2(11):902-910.
De Boeck K, Kent L, Davies J, et al. CFTR biomarkers: time for promotion to surrogate end-point. Eur Respir J. 2013;41(1):203-216.
Bell SC, De Boeck K, Amaral MD. New pharmacological approaches for cystic fibrosis: promises, progress, pitfalls. Pharmacol Ther. 2015;145:19-34.
Ferreira VFC, Oliveira BL, Correia JD G, Santos I, Farinha CM & Mendes FF (2014). Cystic Fibrosis: new molecular imaging tools. Paper presented at the EuroBIC 12-12th European Biological Inorganic Chemistry Conference, Zurich, Switzerland.
Ferreira VFC, Oliveira BL, Santos JD, Correia JDG, Farinha CM, Mendes F. Targeting of the cystic fibrosis transmembrane conductance regulator (CFTR) protein with a technetium-99m imaging probe. ChemMedChem. 2018;13(14):1469-1478.
Freise AC, Wu AM. In vivo imaging with antibodies and engineered fragments. Mol Immunol. 2015;67(2 Pt A):142-152.
Köhler G, Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975;256:495-497.
Smith GP. Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science. 1985;228(4705):1315-1317.
McCafferty J, Griffiths AD, Winter G, Chiswell DJ. Phage antibodies: filamentous phage displaying antibody variable domains. Nature. 1990;348(6301):552-554.
Lowman HB, Bass SH, Simpson N, Wells JA. Selecting high-affinity binding proteins by monovalent phage display. Biochemistry. 1991;30(45):10832-10838.
Barbas CF 3rd, Bain JD, Hoekstra DM, Lerner RA. Semisynthetic combinatorial antibody libraries: a chemical solution to the diversity problem. Proc Natl Acad Sci U S a. 1992;89(10):4457-4461.
Denning GM, Ostedgaard LS, Cheng SH, Smith AE, Welsh MJ. Localization of cystic fibrosis transmembrane conductance regulator in chloride secretory epithelia. J Clin Invest. 1992;89(1):339-349.
Denning GM, Ostedgaard LS, Welsh MJ. Abnormal localization of cystic fibrosis transmembrane conductance regulator in primary cultures of cystic fibrosis airway epithelia. J Cell Biol. 1992;118(3):551-559.
Peters KW, Okiyoneda T, Balch WE, et al. CFTR folding consortium: methods available for studies of CFTR folding and correction. Methods Mol Biol. 2011;742:335-353.
Kaiser E, Colescott RL, Bossinger CD, Cook PI. Color test for detection of free terminal amino groups in the solid-phase synthesis of peptides. Anal Biochem. 1970;34(2):595-598.
Behrendt R, White P, Offer J. Advances in Fmoc solid-phase peptide synthesis. J Pept Sci. 2016;22:4-27.
Miersch S, Sidhu SS. Synthetic antibodies: concepts, potential and practical considerations. Methods. 2012;57:486-498.
Behrendt R, Huber S, Martí R, White P. New t-butyl based aspartate protecting groups preventing aspartimide formation in Fmoc SPPS. J Pept Sci. 2015;21(8):680-687.
Subiros-Funosas R, El-Faham A, Albericio F. Use of Oxyma as pH modulatory agent to be used in the prevention of base-driven side reactions and its effect on 2-chlorotrityl chloride resin. Biopolymers. 2012;98:89-97.
Carmen S, Jermutus L. Concepts in antibody phage display. Brief Funct Genomic Proteomic. 2002;1(2):189-203.
Slama J, Rando RR. The synthesis of glycolipids containing a hydrophilic spacer-group. Carbohydr Res. 1981;88:213-221.
Boumrah D, Campbell MM, Fenner S, Kinsman RG. Spacer molecules in peptide sequences: incorporation into analogues of atrial natriuretic factor. Tetrahedron. 1997;53:6977-6992.
Bartos A, Uray K, Hudecz F. New biotin derivatives for labeling and solubilizing IgG peptides. Biopolymers. 2009;92(2):110-115.
Gardiner J, Mathad RI, Jaun B, Schreiber JV, Flögel O, Seebach D. β-Peptide conjugates: syntheses and CD and NMR investigations of β/α-chimeric peptides, of a DPA-β-decapeptide, and of a PEGylated β-heptapeptide. Helv Chim Acta. 2009;92:2698-2721.
Rothbart SB, Krajewski K, Strahl BD, Fuchs SM. Peptide microarrays to interrogate the “histone code”. Methods Enzymol. 2012;512:107-135.