Korean Journal of Microbiology (미생물학회지)

The Microbiological Society of Korea (한국미생물학회)
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Dna2 Helicase/endonuclease Interacts with a Novel Protein YHR122W Protein in Saccharomyces cerevisiae

Saccharomyces cerevisiae에서 Dna2 helicase/endonuclease와 YHR122W 단백질의 상호작용

  • Lee, Hyun-Sun (Department of Biological Sciences, College of Natural Science, Inha University) ;
  • Choi, Do-Hee (Department of Biological Sciences, College of Natural Science, Inha University) ;
  • Kwon, Sung-Hoon (Department of Biological Sciences, College of Natural Science, Inha University) ;
  • Kim, Na-Yeon (Department of Biological Sciences, College of Natural Science, Inha University) ;
  • Lee, In-Hwan (Department of Biological Sciences, College of Natural Science, Inha University) ;
  • Kim, Hyun-Jung (Department of Biological Sciences, College of Natural Science, Inha University) ;
  • Bae, Sung-Ho (Department of Biological Sciences, College of Natural Science, Inha University)
  • 이현선 (인하대학교 자연과학대학 생명과학과) ;
  • 최도희 (인하대학교 자연과학대학 생명과학과) ;
  • 권성훈 (인하대학교 자연과학대학 생명과학과) ;
  • 김나연 (인하대학교 자연과학대학 생명과학과) ;
  • 이인환 (인하대학교 자연과학대학 생명과학과) ;
  • 김현정 (인하대학교 자연과학대학 생명과학과) ;
  • 배성호 (인하대학교 자연과학대학 생명과학과)
  • Published : 2006.03.01
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Abstract

Saccharomyces cerevisiae Dna2 helicase/endonuclease plays an essential role in removing DNA primers during Okazaki fragment processing in eukaryotic DNA replication. Genome-wide scale co-immunoprecipitation experiments predicted that Dna2 interacts with a novel protein YHR122W (1). In this study, we observed that overexpression of YHR122W gene suppressed the temperature-sensitive phenotype of $dna2\Delta405N$ mutation. To investigate direct interaction between these two proteins, a histidine-tagged recombinant YHR122W protein was expressed and purified from E. coli. Physical interaction between the purified YHR122W and Dna2 proteins was detected by enzyme-linked immunosorbent assays. Further more, the complex formation was most efficient at physiological salt concentration, 150 mM NaCl. The genetic and physical interactions between YHR122W and Dna2 shown in this study suggest that the biological functions of these two proteins may be closely related each other.

Saccharomyces cerevisiae Dna2 helicase/endonuclease는 진핵세포 DNA 복제과정의 Okazaki fragment processing에서 RNA primer를 제거하는데 필수적인 역할을 한다. Genome-wide scale의 면역침전 실험결과, 기능이 알려져 있지 않은 단백질인 YHR122W가 Dna2 단백질과 상호작용한다고 예측되었다 (1). 본 연구에서는 이를 확인하기 위하여 YHR122W 유전자를 효모에서 과량발현시킨 결과, $dna2\Delta405N$ 돌연변이의 온도감수성 표현형이 억제되는 유전학적 상호작용을 관찰하였다. YHR122W 단백질이 Dna2 단백질과 직접적인 삼호작용을 하는지 확인하기 위하여 YHR122W를 대장균에서 재조합 단백질로 발현시키고 단백질을 정제하였다. Enzyme-linked immunosorbent assay를 통한 분석에서 YHR122W 단백질과 Dna2 단백질 사이의 상호작용을 확인하였다. 뿐만 아니라 YHR122W-Dna2 상호작용은 생리적 염도인 150 mM NaCl농도에서 가장 강한 결합을 보였다. 이러한 유전학적 상호작용과 물리적인 상호작용은 YHR122W가 생체내에서 Dna2의 기능과 밀접한 연관이 있을 가능성을 제시하고 있다.

Keywords

References

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