Solution structure of a llama single-domain antibody with hydrophobic residues typical of the VH/VL interface.
The three-dimensional structure of a llama single-domain antibody BrucD4-4 was established by use of solution NMR spectroscopy. BrucD4-4 has Val, Gly, Leu, and Trp residues at positions 37, 44, 45, and 47, which are considered to be a hallmark to distinguish llama VH from V(H)H fragments at the germline level. In contrast to the murine and human VHs, BrucD4-4 has sufficient solubility, is monomeric in solution, and displays high-quality NMR spectra characteristic of well-structured proteins. Amide proton/deuterium exchange and the (15)N relaxation data showed that BrucD4-4 has a classic protein structure with a well-packed core and comparatively mobile surface loops. The three-dimensional architecture of BrucD4-4 is analogous to that of VHs from murine and human F(v)s and camelid V(H)Hs with two pleated beta-sheets formed by four and five beta-strands. A canonical and undistorted beta-barrel exposes a number of hydrophobic residues into the solvent on the surface of the three-dimensional structure. The eight-residue H3 loop folds over the side chain of Val37 similarly to that in llama V(H)Hs; however, this interaction may be transient due to the H3 conformational flexibility. Overall, the surface characteristics of BrucD4-4 with respect to hydrophobicity appear to lie between the human VH domain from Fv Pot and the llama V(H)H fragment HC-V, which may explain its enhanced solubility allowing NMR structural analysis.
Vranken
Wim
W
Biomolecular NMR Laboratory, Biotechnology Research Institute, National Research Council of Canada, 6100 Royalmount Avenue, Montreal, Quebec, Canada H4P 2R2.
Tolkatchev
Dmitri
D
Xu
Ping
P
Tanha
Jamshid
J
Chen
Zhigang
Z
Narang
Saran
S
Ni
Feng
F
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002161:Camelids, New World; D015723:Gene Library; D007128:Immunoglobulin Fragments / Q000737:chemistry*; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry*; D007135:Immunoglobulin Variable Region / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012996:Solutions; D013816:Thermodynamics
D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D012996:Solutions