Purification and characterization of a novel hyaluronan-binding protein (PHBP) from human plasma: it has three EGF, a kringle and a serine protease domain, similar to hepatocyte growth factor activator.
A novel hyaluronan-binding protein (PHBP) was purified from human plasma by affinity chromatography on hyaluronan-conjugated Sepharose. The contaminating IgM and albumin in the partially purified preparation were removed with anti-IgG antibody-conjugated Sepharose and anti-albumin antibody-conjugated Sepharose, respectively, and no other contaminant was observed. Finally, 800 micrograms of PHBP was isolated from 500 ml of human plasma. PHBP gave a single 70-kDa band on SDS-PAGE under non-reducing conditions, and 50-kDa and 17-kDa bands under reducing conditions. Thus, PHBP was a heterodimer composed of 50-kDa and 17-kDa subunits, bridged by a disulfide linkage. Both subunits had novel N-terminal amino acid sequences, indicating that PHBP was a novel hyaluronan-binding protein in human plasma. The amino acid sequence deduced from the nucleotide sequence of the cloned PHBP cDNA exhibited significant homology to that of hepatocyte growth factor activator (HGFA). The results of Northern blot analysis indicated that liver, kidney, and pancreas expressed PHBP mRNA. The predicted structure of PHBP showed three epidermal growth factor (EGF) domains, a kringle domain and a serine protease domain, from its N-terminus, although HGFA has a fibronectin type II domain, an EGF domain, a fibronectin type I domain, an EGF domain, a kringle domain, and a serine protease domain, from its N-terminus.
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PHBP PHBP epidermal growth factor (EGF) domains kringle domain serine protease domain HGFA fibronectin type II domain EGF domain fibronectin type I domain EGF domain serine kringle domain serine protease domain IgM albumin IgG albumin PHBP PHBP PHBP PHBP N PHBP hyaluronan-binding protein PHBP hepatocyte growth factor activator HGFA serine protease domain hyaluronan-binding protein PHBP serine N disulfide N-terminal amino acid amino acid nucleotide serine