Methods for purification of arylsulfatase A
View Patent ↗Methods of producing arylsulfatase A are described herein. The methods can include one or more steps of chromatography. Exemplary chromatographic steps include ion exchange chromatography, mixed mode chromatography, and hydrophobic interaction chromatography. Compositions containing and methods using arylsulfatase A produced by the methods described herein are also disclosed.
1. A method of purifying recombinant arylsulfatase A (ASA) protein, comprising
providing a sample of recombinant ASA protein;
subjecting the sample to a first chromatography step which is anion exchange chromatography,
subjecting the sample from the anion exchange chromatography to a second chromatography step which is mixed-mode chromatography,
subjecting the sample from the mixed-mode chromatography to a third chromatography step which is hydrophobic interaction chromatography, and
subjecting the sample from the hydrophobic interaction chromatography to a fourth chromatography step which is cation exchange chromatography,
wherein fractions from each chromatography step containing recombinant ASA protein are determined by detecting the presence of recombinant ASA protein or enzymatic activity, and
wherein the purified recombinant ASA protein has a specific activity of at least 75 U/mg,
wherein 1 Unit of enzyme activity is defined as the hydrolysis of 1 μmol p-Nitrocatechol Sulphate (pNCS) per minute at 37° C., pH 5.0.
2. The method of claim 1 , wherein the sample of recombinant ASA protein is obtained from cultured cell extract or cell culture supernatant.
3. The method of claim 1 , wherein the cation exchange chromatography is sulfopropyl (SP) cation exchange chromatography.
4. The method of claim 1 , wherein the mixed-mode chromatography is hydroxyapatite (HA) chromatography.
5. The method of claim 1 , wherein the hydrophobic interaction chromatography is phenyl chromatography.