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Purification and properties of dentinal fluid transport stimulating substance from bovine parotid glands.

A substance that stimulates dentinal fluid transport (DFT) through the dentinal tubules in the molar teeth of rats was found in bovine parotid glands. This DFT-stimulating substance was purified by aqueous extraction, isoelectric precipitation, ultrafiltration, gel filtration on Sephadex G-75 and chromatofocusing. By these procedures, 37 mg of purified preparation was obtained from 1 kg of bovine parotid gland. The purified preparation showed a single band on both disc polyacrylamide gel electrophoresis and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. A dose of 10 μg of the purified DFT-stimulating substance per kg of body weight was positive in the bioassay for DFT-stimulating activity, as indicated by the migration of a fluorescent dye through the dentinal tubules in the molar teeth of rats.The molecular weight of DFT-stimulating substance was estimated to be 30000 by both gel filtration on Sephadex G-75 and SDS-polyacrylamide gel electrophoresis. The isoelectric point was found to be pH 6.0 by isoelectric focusing. This active substance was composed of a total of 263 amino acid residues per molecule, had a maximum absorption wavelength (λmax) of 280 nm and an extinction coefficient (E1%1cm) at 280 nm of 15.4, contained less than 1% carbohydrate, and appeared to consist of a single polypeptide chain.