Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Cloning, Expression, and Characterization of a Novel GH-16 β-Agarase from Agarivorans sp. JA-1

Agarivorans sp. JA-1 유래 신규 GH-16 β-agarase의 클로닝, 발현 및 특성

  • Jeon, Myong Je (Department of Pharmaceutical Engineering, College of Medical Life Sciences, Silla University) ;
  • Kim, A-Ram (Department of Pharmaceutical Engineering, College of Medical Life Sciences, Silla University) ;
  • Lee, Dong-Geun (Department of Pharmaceutical Engineering, College of Medical Life Sciences, Silla University) ;
  • Lee, Sang-Hyeon (Department of Pharmaceutical Engineering, College of Medical Life Sciences, Silla University)
  • 전명제 (신라대학교 의생명과학대학 제약공학과) ;
  • 김아람 (신라대학교 의생명과학대학 제약공학과) ;
  • 이동근 (신라대학교 의생명과학대학 제약공학과) ;
  • 이상현 (신라대학교 의생명과학대학 제약공학과)
  • Received : 2012.10.23
  • Accepted : 2012.11.14
  • Published : 2012.11.30
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Abstract

Authors report the glycoside hydrolase (GH) family 16 ${\beta}$-agarase from the strain of Agarivorans sp. JA-1, which authors previously stated as recombinant expression and characterization of GH-50 and GH-118 ${\beta}$-agarase. It comprised an open reading frame of 1,362 base pairs, which encodes a protein of 49,830 daltons consisting of 453 amino acid residues. Valuation of the total sequence showed that the enzyme has 98% nucleotide and 99% amino acid sequence similarities to those of GH-16 ${\beta}$-agarase from Pseudoalteromonas sp. CY24. The gene corresponding to a mature protein of 429 amino acids was recombinantly expressed in Escherichia coli, and the enzyme was purified to homogeneity by affinity chromatography. It showed maximal activity at $40^{\circ}C$ and pH 5.0, representing 67.6 units/mg. Thin layer chromatography revealed that mainly neoagarohexaose and neoagarotetraose were produced from agarose. The enzyme would be valuable for the industrial production of functional neoagarooligosaccharides.

이전 연구에서 저자들이 Glycoside hydrolase family 50 (GH-50)과 GH-118 ${\beta}$-agarase들의 발현과 특성을 보고한 Agarivorans sp. JA-1 균주로부터 신규의 GH-16 ${\beta}$-agarase를 보고하고자 한다. 본 유전자는 1,362 염기쌍으로 구성되어 있으며, 453 아미노산 잔기로 구성된 49,830 Da의 단백질을 암호화한다. 본 효소는 Pseudoalteromonas sp. CY24 유래의 GH-16 ${\beta}$-agarase와 98%의 염기서열 상동성과 99%의 아미노산서열 상동성을 나타냈다. 신호서열을 제외한 429 아미노산으로 구성된 성숙단백질에 해당하는 유전자를 E. coli BL21 (DE3) 세포에서 재조합 발현시킨 후, 친화성 크로마토그래피로 효소를 정제하였다. 정제된 효소는 $40^{\circ}C$와 pH 5.0에서 67.6 U/mg의 최적 활성을 보였다. 아가로스를 기질로 한 효소분해산물의 박막크로마토그래피 분석결과, neoagarohexaose와 neoagarotetraose가 주산물로 생산되는 것을 알 수 있었다. 본 효소는 기능성 한천올리고당의 산업적 생산에 활용 가능할 것으로 기대된다.

Keywords

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