Purification, characterization, and relation to bikunin of rat urinary trypsin inhibitors

Two forms of urinary trypsin inhibitor (UTI-1 and UTI-2) were purified from pooled urine of normal male rats to apparent homogeneity by salting out, affinity chromatography, gel filtration, and reverse-phase HPLC. UTIs-1 and 2 were shown to be thermostable glycoproteins with the respective molecular weights of 22,000 and 18,000 estimated by SDS-PAGE. These inhibitors combined with bovine trypsin in a 1:1 molar ratio: the K_d values were 2.5 × 10^–10 and 2.3 × 10^–10 M, respectively. Amino acid composition and sequence analysis indicated that UTI-1 corresponded to rat bikunin of which the amino acid sequence was deduced from a rat liver cDNA clone encoding ₁-microglobulin [Lindqvist et al. (1992), Biochim. Biophys. Acta1130, 63–67] except that the protein sequence seemed to lack C-terminal serine, and UTI-2 corresponded to UTI-1 lacking N-terminal 21 amino acid residues.