Isolation of three forms of cystatin from submandibular saliva of isoproterenol-treated rats, its properties and kinetic data.
Three rat salivary cystatins (designated as RSC-1, RSC-2 and RSC-3) induced by chronic isoproterenol (IPR) treatment, but not detected in normal rats, were purified from submandibular saliva of chronically IPR-treated rats by Mono-Q, hydroxyapatite and TSKgel Phenyl-5PW chromatographies. Their molecular weights (Mr) and isoelectric points (pI) differed from each other as follows: RSC-1 (Mr 16,500, pI 4.4), RSC-2 (Mr 15,500, pI 4.4) and RSC-3 (Mr 14,500, pI 4.5). The amino acid compositions of these inhibitors were very similar and the three forms showed complete immunological identity in a double immunodiffusion system. The partial amino acid sequence results showed that these inhibitors belonged to family 2 of the cystatin superfamily. These three forms strongly inhibited the enzyme activities of ficin and papain, but not of cathepsin B and trypsin. The inhibition constants (Ki) of RSC-1, RSC-2 and RSC-3 for ficin were 0.19, 0.50 and 0.012 nM, and for papain were 1.5, 0.93 and 0.03 nM, respectively. RSC-3 inhibited ficin and papain more strongly than did RSC-1 and RSC-2.
Nishiura
T
T
Department of Oral Biochemistry, Fukuoka Dental College, Japan.
Ishibashi
K
K
Abe
K
K
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D015891:Cystatins / Q000032:analysis / Q000302:isolation & purification* / Q000494:pharmacology; D005779:Immunodiffusion; D007545:Isoproterenol / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D051381:Rats; D011919:Rats, Inbred Strains; D012463:Saliva / Q000737:chemistry*; D013363:Submandibular Gland / Q000737:chemistry*
D000596:Amino Acids; D015891:Cystatins; D007545:Isoproterenol