Disulfide Bond Introduction for General Stabilization of Immunoglobulin Heavy-Chain Variable Domains

Several antibody fragment engineering techniques aim at intrinsic stability enhancement, but are not applied in a truly generic way. Here, a strategy is proposed whereby consistent gain in stability is accomplished by introducing a specific disulfide bond between two opposite β-strands in the hydrop...

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Bibliographic Details
Published inJournal of molecular biology Vol. 377; no. 2; pp. 478 - 488
Main Authors Saerens, Dirk, Conrath, Katja, Govaert, Jochen, Muyldermans, Serge
Format Journal Article
LanguageEnglish
Published England Elsevier Ltd 21.03.2008
Subjects
Amino Acid Sequence
antibody
Cysteine - genetics
Cysteine - metabolism
cystine
Disulfides - chemistry
Disulfides - metabolism
Immunoglobulin Heavy Chains - chemistry
Immunoglobulin Heavy Chains - classification
Immunoglobulin Heavy Chains - genetics
Immunoglobulin Heavy Chains - metabolism
Immunoglobulin Variable Region - chemistry
Immunoglobulin Variable Region - classification
Immunoglobulin Variable Region - genetics
Immunoglobulin Variable Region - metabolism
Models, Molecular
Molecular Sequence Data
Mutation - genetics
Nanobody
Nanostructures - chemistry
Protein Folding
Protein Structure, Tertiary
Sequence Alignment
stability
β-strand
CDR
Fv
Nanobody
VH
IMGT
antibody
β-strand
hPSA
cystine
GdmHCl
stability
PDB
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Summary:Several antibody fragment engineering techniques aim at intrinsic stability enhancement, but are not applied in a truly generic way. Here, a strategy is proposed whereby consistent gain in stability is accomplished by introducing a specific disulfide bond between two opposite β-strands in the hydrophobic core of the immunoglobulin heavy-chain variable domain of heavy-chain antibodies (Nanobody). Besides the rational design of a disulfide bond between residues 39 and 87, a Nanobody harboring an extra naturally occurring cystine between residues 54 and 78 was compared to an equivalent Nanobody without that cystine. Both novel disulfide cross-links were introduced in several Nanobodies in various combinations. Interestingly, only the extra naturally occurring cystine consistently increased the conformational and thermal stabilities of wild-type Nanobodies without affecting antigen binding.
Bibliography:ObjectType-Article-1
SourceType-Scholarly Journals-1
ObjectType-Feature-2
content type line 23
ISSN:0022-2836
1089-8638
DOI:10.1016/j.jmb.2008.01.022