Cloning, expression, and crystallization of the V delta domain of a human gamma delta T-cell receptor.

Printer-friendly versionPrinter-friendly versionPDF versionPDF version
TitleCloning, expression, and crystallization of the V delta domain of a human gamma delta T-cell receptor.
Publication TypeJournal Article
Year of Publication1996
AuthorsLebedeva, MI, Fields, BA, Spits, H, Panchamoorthy, G, Brenner, MB, Mariuzza, RA
JournalProtein Sci
Date Published1996 Dec
KeywordsAmino Acid Sequence, Base Sequence, Cloning, Molecular, Crystallization, Crystallography, X-Ray, Escherichia coli, Gene Expression, Humans, Immunoglobulin Variable Region, Molecular Sequence Data, Receptors, Antigen, T-Cell, gamma-delta

T-lymphocytes recognize a wide variety of antigens through highly diverse cell-surface glycoproteins known as T-cell receptors (TCRs). These disulfide-linked heterodimers are composed of alpha and beta or gamma and delta polypeptide chains consisting of variable (V) and constant (C) domains non-covalently associated with at least four invariant chains to form the TCR-CD3 complex. It is well established that alpha beta TCRs recognize antigen in the form of peptides bound to molecules of the major histocompatibility complex (MHC); furthermore, information on the three-dimensional structure of alpha beta TCRs has recently become available through X-ray crystallography. In contrast, the antigen specificity of gamma delta TCRs is much less well understood and their three-dimensional structure is unknown. We have cloned the delta chain of a human TCR specific for the MHC class I HLA-A2 molecule and expressed the V domain as a secreted protein in the periplasmic space of Escherichia coli. Following affinity purification using a nickel chelate adsorbent, the recombinant V delta domain was crystallized in a form suitable for X-ray diffraction analysis. The crystals are orthorhombic, space group P2(1)2(1)2 with unit cell dimensions a = 69.9, b = 49.0, c = 61.6 A. and diffract to beyond 2.3 A resolution. The ability of a V delta domain produced in bacteria to form well-ordered crystals strongly suggests that the periplasmic space can provide a suitable environment for the correct in vivo folding of gamma delta TCRs.

Alternate JournalProtein Sci.
PubMed ID8976572
PubMed Central IDPMC2143324