Characterization of Human Interferon-γ Receptor Purified from Placenta

Abstract

Membranes prepared from human placenta were used for characterization of the receptor for human interferon-γ (HuIFN-γ) after large-scale purification. HuIFN-γ linked covalently to Affigel-10 was used for the purification of the receptor from octylglucoside-solubilized placental membranes. Radiolabeled IFN-γ [³²P]HuIFN-γ, was used in binding and cross-linking studies to detect the receptor at different stages of the purification. From binding assays it was calculated that an average placenta contained 90–120 μg of receptor with a K _d value of 1.3 × 10⁻⁹ M. Thus, human placenta is a rich and convenient source of receptor for IFN- γ. When purified receptor was cross-linked to [³²P]HuIFN-γ, a variety of cross-linked bands were detected dependent on the preparation conditions. The use of protease inhibitors in the course of processing prevented degradation of the 90-kD intact receptor, showing that the lower-molecular-weight products detected in previous studies are degradation products of the receptor. Furthermore, a 20-kD fragment of the receptor was found to be active in binding HuIFN-γ.