TY - JOUR
T1 - Preparation and crystallization of the disulfide-linked HLA-G dimer
AU - Shiroishi, Mitsunori
AU - Kohda, Daisuke
AU - Maenaka, Katsumi
N1 - Funding Information:
We thank K. Hasegawa for helping data collection at Spring-8 and S. Kollnberger for critical reading. MS was supported in part by Sasakawa Scientific Research Grant from the Japan Science Society and by JSPS Research Fellowship for Young Scientist. KM and DK were supported in part by Ministry of Education, Science, Sports, Culture and Technology of Japan, and the Protein 3000 project. KM was supported in part by Kanae foundation.
PY - 2006/5
Y1 - 2006/5
N2 - HLA-G is a non-classical MHC class I, which binds to inhibitory receptors, such as Leukocyte Ig-like receptors, to induce a wide range of tolerogenic immunological effects. HLA-G can be expressed as a disulfide-liked dimer both in solution and at the cell surface. However, the three-dimensional structure of the HLA-G dimer is unknown. Here, we report the crystallization of the disulfide-linked dimer form of HLA-G by adding dithiothreitol (DTT), enabling a 3.2-Å data set to be collected. We also show that DTT promotes disulfide bond exchange of refolded HLA-G, whose free cysteine was protected, thus facilitating its dimerization. This technique could also be applied for disulfide-mediated dimer/multimer formation of refolded proteins harbouring free cysteines.
AB - HLA-G is a non-classical MHC class I, which binds to inhibitory receptors, such as Leukocyte Ig-like receptors, to induce a wide range of tolerogenic immunological effects. HLA-G can be expressed as a disulfide-liked dimer both in solution and at the cell surface. However, the three-dimensional structure of the HLA-G dimer is unknown. Here, we report the crystallization of the disulfide-linked dimer form of HLA-G by adding dithiothreitol (DTT), enabling a 3.2-Å data set to be collected. We also show that DTT promotes disulfide bond exchange of refolded HLA-G, whose free cysteine was protected, thus facilitating its dimerization. This technique could also be applied for disulfide-mediated dimer/multimer formation of refolded proteins harbouring free cysteines.
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U2 - 10.1016/j.bbapap.2005.10.006
DO - 10.1016/j.bbapap.2005.10.006
M3 - Article
C2 - 16290109
AN - SCOPUS:33646510515
VL - 1764
SP - 985
EP - 988
JO - Biochimica et Biophysica Acta - Proteins and Proteomics
JF - Biochimica et Biophysica Acta - Proteins and Proteomics
SN - 1570-9639
IS - 5
ER -