Reference : Identification of a universal VHH framework to graft non-canonical antigen-binding loops...
Scientific journals : Article
Life sciences : Biochemistry, biophysics & molecular biology
http://hdl.handle.net/2268/22340
Identification of a universal VHH framework to graft non-canonical antigen-binding loops of camel single-domain antibodies
English
Saerens, Dirk [ > > ]
Pellis, Mireille [ > > ]
Loris, Remy [ > > ]
Pardon, Els [ > > ]
Dumoulin, Mireille mailto [Université de Liège - ULg > Département des sciences de la vie > Laboratoire d'Enzymologie et Repliement des Protéines, Centre d'Ingénierie des Protéines > >]
Matagne, André mailto [Université de Liège - ULg > Département des sciences de la vie > Laboratoire d'Enzymologie et Repliement des Protéines, Centre d'Ingénierie des Protéines > >]
Wyns, Lode [ > > ]
Muyldermans, Serge [ > > ]
2005
Journal of Molecular Biology
Academic Press
352
597-607
Yes (verified by ORBi)
International
0022-2836
1089-8638
London
United Kingdom
[en] nanobodies ; camelid antibodies ; protein stability ; folding ; thermodynamics ; affinity ; CDR grafting ; VHH ; generic framework
[en] Camel single-domain antibody fragments (VHHs) are promising tools in
numerous biotechnological and medical applications. However, some
conditions under which antibodies are used are so demanding that they
can be met by only the most robust VHHs. A universal framework offering
the required properties for use in various applications (e.g. as intrabody, as
probe in biosensors or on micro-arrays) is highly valuable and might be
further implemented when employment of VHHs in human therapy is
envisaged. We identified the VHH framework of cAbBCII10 as a potential
candidate, useful for the exchange of antigen specificities by complementarity
determining region (CDR) grafting. Due to the large number of CDRH
loop structures present on VHHs, this grafting technique was expected
to be rather unpredictable. Nonetheless, the plasticity of the cAbBCII10
framework allows successful transfer of antigen specificity from donor
VHHs onto its scaffold. The cAbBCII10 was chosen essentially for its high
level of stability (47 kJ/mol), good expression level (5 mg/l in E. coli)
and its ability to be functional in the absence of the conserved disulfide
bond. All five chimeras generated by grafting CDR-Hs, from donor VHHs
belonging to subfamily 2 that encompass 75% of all antigen-specific VHHs,
on the framework of cAbBCII10 were functional and generally had an
increased thermodynamic stability. The grafting of CDR-H loops from
VHHs belonging to other subfamilies resulted in chimeras of reduced
antigen-binding capacity.
http://hdl.handle.net/2268/22340

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