Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Characterization of Ferritin Isolated from Dog Spleen

개의 비장에서 분리한 페리틴의 특성

  • Park Jae-Hag (Laboratory of Immunobiology, Department of Microbiology, College of Natural Sciences, Kyungpook National University) ;
  • Jun Do Youn (Institute of Genetic Engineering, Kyungpook National University) ;
  • Kim Young Ho (Laboratory of Immunobiology, Department of Microbiology, College of Natural Sciences, Kyungpook National University)
  • 박재학 (경북대학교 자연과학대학 미생물학과 면역생물학연구실) ;
  • 전도연 (경북대학교 유전공학연구소) ;
  • 김영호 (경북대학교 자연과학대학 미생물학과 면역생물학연구실)
  • Published : 2005.06.01
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Abstract

Ferritin is known to be the principle iron-storage protein in a wide variety of rganisms. The electro­phoretic mobility and immunological cross-reactivity of dog splenic ferritin were compared with those of horse, bovine, and pig splenic ferritin after isolation using heat treatment, salting out, column chromatography, and ultrafiltration. These isolation methods allowed the recovery of $\~84{\mu}g$ of the ferritin per g of spleen. The iron content in the dog ferritin was $22.7\%$, which appeared to be higher than those in the other mammalian ferritins tested. The electrophoretic mobility of the dog ferritin under nondenaturing conditions was similar to its bovine counterpart, whereas it was more identical to pig and horse ferritins on an SDS-polyacrylamide gel. The molecular weight of the dog ferritin subunit was 19.5 kDa on an SDS-polyacrylarnide gel, and the subunit was unable to bind with iron. The polyclonal anti-dog ferritin raised in rats was able to cross-react with the pig, bovine, and horse ferritins, upon Ouchterlony double immunodiffusiion. A Western blot analysis also revealed that the anti-dog ferritin, which specifically bound with the dog ferritin subunit, could also recognize the horse, bovine, and pig ferritin subunits and the maximum cross-reactivity was exhibited with the pig ferritin subunit, indicating that the dog ferritin is immunochemically more similar to the pig ferritin than its other mammalian counterparts. Accordingly, these results elucidate the biochemical and immunochemical characteristics of dog ferritin that might have a potential to be applied as an oral iron supplement to treat iron deficiency anemia.

페리틴은 생체 내 주요 철 저장 단백질로서 포유류에서 세균류에 이르기까지 다양한 생명체에 존재한다. 페리틴 분자는 $18\~22 kDa$의 단량체 24개가 결합된 약 240 kDa분자량을 지닌 거대분자이다. 본 연구에서는 개의 비장에서부터 열처리, 염석, 컬럼 크로마토그래피, 그리고 한외여과 등의 방법으로 페리틴을 정제한 후, 그 전기영동상의 특성 및 면역화학적 특성을 말, 소, 돼지 등의 비장 유래 페리틴과 비교 분석하였다. 이러한 정제방법에 의해 개의 비장으로부터 페리틴의 양은 비장 1g당 약$84{\mu}g$이었다. 정제된 개 페리틴의 철 함량은 $22.7\%$로서 함께 비교 검토한 다른 동물 유래의 페리틴에 비해 가장 높게 나타났다. 개 페리틴의 비변성 겔에서의 이동상은 소페리틴과 유사하였고, 변성 겔에서는 돼지 및 말의 페리틴과 유사하였다. SDS-PACE상에서 나타난 개 페리틴 subunit의 분자량은 19.5kDa이였으며 이때 SDS-PACE상의 ferritin subunit은 철과 결합하지 않는 것으로 확인되었다. 개 페리틴의 면역화학적 특성을 다른 동물유래의 페리틴과 비교하고자, 개의 페리틴에 대한 다클론 항체를 쥐에서 생산하였다. 생산된 항-페리틴 항체를 이용하여 Ouchterlony doulbe immunodiffusion방법으로 개, 소, 말, 돼지 페리틴들을 항원으로 항원-항체반응을 조사한 결과, 항-개 페리틴 항체가 소, 말, 돼지의 페리틴과도 항원-항체반응을 일으킬 수 있는 것으로 나타났다. 이는 개, 소, 말, 돼지 페리틴들이 항원적 동일성을 지니고 있음을 시사한다. 항-개 페리틴 항체를 이용하여 타 동물 유래의 페리틴에 대한 Western blot analysis를 시도한 결과, 항-개 페리틴 항체가 개 페리틴 및 돼지 폐리틴에는 강하게 반응하였으나 말과 소 유래의 페리틴에 대해서는 비교적 약하게 반응하여, 개의 페리틴이 면역화학적으로 돼지 페리틴과 가장 유사한 것으로 나타났다. 이상의 결과들은 개의 페리틴에 대한 생화학적 및 면역학적 특성을 제시한다.

Keywords

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