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Solution structure and backbone dynamics of an antigen‐free heavy chain variable domain (VHH) from Llama
Author(s) -
Renisio JeanGuillaume,
Pérez Janice,
Czisch Michael,
Guenneugues Marc,
Bornet Olivier,
Frenken Leon,
Cambillau Christian,
Darbon Hervé
Publication year - 2002
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.10096
Subject(s) - heteronuclear molecule , chemistry , single domain antibody , antiparallel (mathematics) , complementarity determining region , nuclear magnetic resonance spectroscopy , crystallography , biophysics , nuclear magnetic resonance , stereochemistry , antibody , physics , peptide sequence , biochemistry , biology , quantum mechanics , magnetic field , immunology , gene
Camelids, (dromedaries, camels, and llamas) produce heavy‐chains antibodies, with their antigen recognition sites composed of a single VH‐like domain, referred to as VHH. The solution structure of one of these VHHs domains (VHH‐H14), raised against the α subunit of the human chorionic gonadotropin hormone (hCG), has been determined by 15 N heteronuclear three‐dimensional NMR spectroscopy. The framework is well resolved within the set of 20 best‐calculated NMR structures and is close to that of classical VH domains from vertebrate antibodies, consisting of two antiparallel β‐sheets organized in a β‐barrel. Loops display a lower precision, especially the Complementarity Determining Regions (CDRs), involved in antigen recognition. Comparison of the three‐dimensional VHH‐H14 solution structure with its previously solved crystal structure (Spinelli et al., Nature Struct. Biol. 1996;3:752–757) reveals a high similarity to the framework, whereas significant conformational differences occur on CDRs, leading to the assumption that the antigen recognition site is a more mobile part. In order to deepen our insights into the dynamics of VHH‐H14 in solution, 15 N relaxation was measured with longitudinal R1 and transverse R2 self‐relaxation rates, and 15 N steady‐state heteronuclear nuclear Overhauser enhancements (NOE), making it possible to probe picosecond‐to‐millisecond internal motions. Determination of dynamic parameters (S 2 , τ e , and Rex) through the Lipari‐Szabo Model‐free approach enables the identification of several regions with enhanced dynamics. Especially, the mobility measurements from NMR confirm that the antigen recognition site is the most mobile part of the VHH‐H14 domain on picosecond‐to‐nanosecond fast time scales. Several residues belonging to the three CDRs are submitted to chemical exchange processes occurring on slow microsecond‐to‐millisecond time scales, suggesting that the formation of the VHH/antigen complex should be accompanied by structural changes. Proteins 2002;47:546–555. © 2002 Wiley‐Liss, Inc.