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Purification and crystallization of a complex between human interferon γ receptor (extracellular domain) and human interferon γ
Author(s) -
Windsor William T.,
Walter Leigh J.,
Syto Rosalinda,
Fossetta James,
Cook William J.,
Nagabhushan Tattanahalli L.,
Walter Mark R.
Publication year - 1996
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/(sici)1097-0134(199609)26:1<108::aid-prot10>3.0.co;2-k
Subject(s) - chemistry , crystallization , interferon , crystallography , receptor , biochemistry , biology , organic chemistry , genetics
X‐ray diffraction quality crystals have been obtained from a complex between interferon γ and the extracellular domain of its high‐affinity cell surface receptor. The crystals were obtained from interferon γ/interferon γ receptor complexes purified by size exclusion chromatography. Diffraction quality crystals required analyzing these complex samples by isoelectric focusing gels to select purified complex fractions devoid of unbound interferon γ. These studies used interferon γ receptor engineered with an eight amino acid N‐terminal deletion to eliminate heterogeneity generated due to proteolytic cleavage. In addition, the receptor was expressed in an E. coli secretion cell line which eliminated the need to refold the protein. Hexagonal crystals were grown from 1.6 M ammonium phosphate solutions and belong to a spacegroup of P6 5 22 with unit cell dimensions a = 145.9 Å and c = 180.3 Å. These crystals diffract to at least 2.9 Å resolution when exposed to synchrotron radiation. SDS PAGE analysis of the crystals demonstrated that both interferon γ and the receptor were present. Analysis of the x‐ray diffraction data revealed that the crystals contain complexes with a stoichiometry of 2:1 receptor: ligand within the crystallographic asymmetric unit and consist of approximately 55% solvent. © 1996 Wiley‐Liss, Inc.