Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Cloning and Characterization of a Novel Carboxylesterase Gene from Cow Rumen Metagenomic Library

소 반추위 메타게놈에서 새로운 carboxylesterase 유전자 클로닝 및 유전산물의 특성

  • Asraful Islam, Shah Md. (Division of Applied Life Science (BK21 Program), Gyeongsang National University) ;
  • Kim, Min-Keun (Gyeongsangnam-do Agricultural Research and Extension Service) ;
  • Renukaradhya, K. Math (Division of Applied Life Science (BK21 Program), Gyeongsang National University) ;
  • Srinivasa, Reddy R.N. (Division of Applied Life Science (BK21 Program), Gyeongsang National University) ;
  • Kim, Eun-Jin (Division of Applied Life Science (BK21 Program), Gyeongsang National University) ;
  • Kim, Jung-Ho (Department of Agricultural Chemistry, Sunchon National University) ;
  • Kim, Hoon (Department of Agricultural Chemistry, Sunchon National University) ;
  • Yun, Han-Dae (Division of Applied Life Science (BK21 Program), Gyeongsang National University)
  • Received : 2010.07.23
  • Accepted : 2010.08.10
  • Published : 2010.09.30
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Abstract

The gene encoding esterase enzyme was cloned from a metagenomic library of cow rumen bacteria. The esterase gene (est1R) was 2,465 bp in length, encoding a protein of 366 amino acid residues, and the molecular weight of the enzyme was 61,166 Da. Est1R of rumen cosmid library shared 5.9% amino acid identity with Est1R (P37967) of PNB carboxylesterase, 6.1% with Est1R (1EEAA) of acetylcholinesterase and 6.1% with Est1R (1H23A) of chain A. BlastP in NCBI database analysis of Est1R revealed that it was not homologous to previous known lipases and esterases. Est1R showed optimum activity at pH 7.0 and $40^{\circ}C$. On the other hand, the enzyme was found to be most active without organic solvent, followed by 95% activity with methanol, and the enzyme activity was highly affected by hexane (lost 51% activity). Therefore, the novel esterase gene est1R is likely obtainable from cow rumen metagenome and may be utilized for industrial purposes.

한우의 반추위에서 게놈 DNA를 분리하여 메타게놈 은행을 구축한 다음 carboxylesterase를 암호화하는 유전자를 클로닝 및 유전자를 선별하였다. 선별된 유전자의 DNA 염기서열 및 아미노산 서열을 분석하고 유전산물의 생화학적인 특성을 조사하였다. est1R 유전자는 2,465 bp로 366개의 아미노산 잔기를 가진 단백질을 암호화하였으며 이 단백질의 이론적인 분자량은 61,166 Da이었다. Est1R단백질은 PNB carboxylesterase (P37967), acetylcholinesterase (1EEAA) 및 chain A (1H23A)와 각각 5.9%, 6.1%, 6.1% 상동성을 가지고 있었다. 이러한 검색 결과 기존의 알려진 lipase 및esterase와의 상동성이 낮은 것으로 보아 새로운 그룹의 효소로 추정된다. Est1R효소의 최적 pH는 7.0 근방이었으며 최적 온도는 $40^{\circ}C$ 부근이었다. 한편 10% 유기용매를 함유한 기질의 효소활성측정에서 대조구에 비해 methanol은 95%의 상대적인 활성을 가진 반면에 hexane 용액에서는 그 활성이 반으로 감소하였다. 따라서 유기용매 농도의 작용성에 따라 이 효소의 산업적 이용성도 가능하리라 추정된다.

Keywords

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