A METHOD FOR PURIFICATION OF RECOMBINANT HUMAN ALPHA-GALACTOSIDASE A FROM MATERIAL CONTAINING CONTAMINANT HOST CELL PROTEINS

• Main Authors: KAWASAKI ATSUKO, SUGIMURA ATSUSHI, FUKUI TSUYOSHI, MIHARA KAZUTOSHI, ITO YAE, HATANO YUKICHI
• Format: Patent
• Language: English; Korean
• Published: 31.08.2017
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS THEREOF; CULTURE MEDIA; ENZYMOLOGY; HUMAN NECESSITIES; HYGIENE; MEDICAL OR VETERINARY SCIENCE; METALLURGY; MICROBIOLOGY; MICROORGANISMS OR ENZYMES; MUTATION OR GENETIC ENGINEERING; PREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; SPIRITS; VINEGAR; WINE

개시되는 것은 고순도로, 대규모로 재조합 인간 α-갈락토시다아제 A (rh α-Gal A) 를 제조하는 방법이다. 상기 방법은 (a) 무혈청 배지에서 rh α-Gal A-생성 포유동물 세포를 배양하는 단계, (b) 배양 상청액을 수집하는 단계, (c) 음이온-교환 컬럼 크로마토그래피에 배양 상청액을 적용하는 단계, (d) 소수성 컬럼 크로마토그래피 단계, (e) 포스페이트 기에 대한 친화성을 갖는 물질을 고체 상으로서 사용하는 컬럼 크로마토그래피 단계, (f) 양이온-교환 컬럼 크로마토그래피 단계, (g) 염료-친화성 컬럼 크로마토그래피 단계, 및 (h) 겔 여과 컬럼 크로마토그래피 단계를 순서대로 포함한다. Disclosed is a method for production of recombinant human alpha-galactosidase A (rh alpha-Gal A) in a large scale, with a high purity. The method comprises the steps of (a) culturing rh alpha-Gal A-producing mammalian cells in a serum-free medium, (b) collecting culture supernatant, (c) subjecting the culture supernatant to anion-exchange column chromatography, (d) to hydrophobic column chromatography, (e) to a column chromatography employing as solid phase a material having affinity for phosphate group, (f) to cation-exchange column chromatography, (g) to dye-affinity column chromatography, and (h) to gel filtration column chromatography, in the order.