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.
PHBP is a novel human plasma hyaluronan-binding protein that shows significant homology in amino acid sequence to hepatocyte growth factor activator. Two overlapping clones that encode the human plasma hyaluronan-binding protein (PHBP) gene (HABP2) were isolated and characterized. The PHBP gene spans 35 kb and is composed of 13 exons from 37 to 1,394 bp in size with consensus splice sites. The gene's regulatory sequences contain putative promoter elements, but no typical TATA box. Some exons of this gene showed significant similarities to those of coagulation factor XII, tissue-type plasminogen activator, and urokinase genes in nucleotide length and in intron phasing. We also report the chromosome mapping of this gene by fluorescence in situ hybridization (FISH) using a genomic DNA fragment as a probe. The PHBP gene (HABP2) was located on chromosome 10q25-q26.
The pig counterpart of human IHRP has been isolated from pig serum, and cDNA clones encoding this counterpart were isolated and characterized. The amino acid sequence of pig IHRP predicted from the nucleotide sequence of its cDNA shows reasonable homology to that of human IHRP. The nucleotide sequence of pig IHRP cDNA is identical to that of the mRNA partially determined to be the heavy chain of pig ITI [Buchman et al. (1990) Surgery 108, 560-566]; it is one of the major mRNAs induced in pig liver on cardiogenic shock. Thus we concluded that the reported mRNA should code for IHRP and not for the heavy chain of ITI. The pig IHRP also seems to be identical to pig-MAP, which was recently reported to be a major acute phase serum protein in pig [Gonzalez-Roman et al. (1995) FEBS Lett. 371, 227-230]. The results suggest that IHRP might be involved in acute phase reactions.
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