US 5,496,718 AGranted

Chondroitinase ABC isolated from proteus vulgaris ATCC 6896

AssigneeSeikagaku Kogyo Kabushiki Kaisha (Seikagaku Corporation)

InventorsNobukazu Hashimoto, Hideo Mochizuki, Akio Hamai

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Abstract

A crystallizable, purified chondroitinase ABC having a molecular weight of about 100,000 dalton by the measurement of the SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and the measurement by the gel permeation chromatography method, having alanine as the N-terminal amino acid and proline as the C-terminal amino acid. A process for the purification of the crystallizable purified chondroitinase ABC comprising removing nucleic acid from an surfactant solution extract obtained from cells of chondroitinase ABC-producing microorganisms and chromatographically treating by concentration gradient elution using a weak cation exchange resin or a strong cation exchange resin. A composition comprising a chondroitinase and serum albumin, gelatin, or a nonionic surfactant. The chondroitinase ABC is isolated from Proteus vulgaris ATCC 6896.

Claims1

Dependent claims

A purified chondroitinase ABC from Proteus vulgaris ATCC 6896 having the following characteristics:

(i) a single polypeptide chain having a molecular weight of about 100,000 daltons as measured by a single band in SDS-polyacrylamide gel electrophoresis (SDS-PAGE), in a reduced or unreduced state, and by gel permeation chromatography,
(ii) an isoelectric point from about 8.2 and about 8.5,
(iii) an optimum pH of from about 8.0 to 8.2,
(iv) an optimum reaction temperature of 37.degree. C.,
(v) a specific activity of 300 U/mg or more, and
(vi) a single peak as determined by HPLC;
wherein activity of the chondroitinase ABC is inhibited by contact with a metal ion selected from the group consisting of Zn.sup.2+, Ni.sup.2+, Fe.sup.3+, and Cu.sup.2+
wherein the purified chondroitinase ABC is essentially free of nucleic acid, endotoxin and protease activity, and has a terminal amino acid sequence of Ala-Thr-X-Asn-Pro-Ala-Phe-Asp-Pro-Ser-Leu-Pro, wherein X is undetermined.

5 more elements

Dependencies are inferred from the claim text.