Journal of Plant Biotechnology

  • 제48권3호
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  • Pages.165-172
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  • 2021
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  • 1229-2818(pISSN)
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  • 2384-1397(eISSN)
한국식물생명공학회 (The Korean Society of Plant Biotechnology)
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벼의 차세대 단백질체 분석을 위한 질량분석기 호환의 광분해성 계면활성제의 적용

Application of mass-spectrometry compatible photocleavable surfactant for next-generation proteomics using rice leaves

  • 신혜원 (부산대학교 식물생명과학과) ;
  • 응웬반쯔엉 (부산대학교 식물생명과학과) ;
  • 정주용 (부산대학교 식물생명과학과) ;
  • 이기현 (부산대학교 식물생명과학과) ;
  • 장정우 (부산대학교 식물생명과학과) ;
  • 윤진미 (부산대학교 식물생명과학과) ;
  • 라비굽타 (국민대학교 교양대학) ;
  • 김선태 (부산대학교 식물생명과학과) ;
  • 민철우 (부산대학교 식물생명과학과)
  • Shin, Hye Won (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Nguyen, Truong Van (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Jung, Ju Young (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Lee, Gi Hyun (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Jang, Jeong Woo (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Yoon, Jinmi (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Gupta, Ravi (College of General Education, Kookmin University) ;
  • Kim, Sun Tae (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University) ;
  • Min, Cheol Woo (Department of Plant Bioscience, Life and Industry Convergence Research Institute, Pusan National University)
  • 투고 : 2021.09.24
  • 심사 : 2021.09.29
  • 발행 : 2021.09.30
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초록

식물과 동물 모두 단백질체 분석을 위해 계면활성제를 사용하여 단백질을 효과적으로 가용화 하는 것은 매우 중요하다. 여러 계면활성제 중 광분해성의 Azo는 MS 분석에 적합하며 단백질을 효과적으로 잘 가용화 하는 등의 여러 이점을 보여주었다. 그러나 대부분이 동물 단백질체 분석에 적용되어 있었으며 식물 단백질체 분석에의 적용은 미비하였다. 따라서 본 연구에서는 벼 잎의 단백질체 분석을 위한 계면활성제로 Azo를 사용하여 SDS와 비교 분석을 수행하였다. 비표지 단백질체 정량 분석, 단백질 기능 분석, 세포내 단백질체의 위치 확인 분석 및 KEGG 경로 분석을 수행하였으며, 그 결과 SDS와 비교하였을 때 Azo의 단백질 가용화 효율이 전혀 떨어지지 않음을 확인하였다. 이는 앞으로 벼 뿐만 아니라 식물단백질체 분석 시에 광분해성 계면활성제인 Azo의 적용 가능성이 무궁무진함을 의미 한다.

The solubilization of isolated proteins into the adequate buffer containing of surfactants is primary step for proteomic analysis. Particularly, sodium dodecyl sulfate (SDS) is the most widely used surfactant, however, it is not compatible with mass spectrometry (MS). Therefore, it must be removed prior to MS analysis through rigorous washing, which eventually results in inevitable protein loss. Recently, photocleavable surfactant, 4-hexylphenylazosulfonate (Azo), was reported which can be easily degraded by UV irradiation and is compatible with MS during proteomic approach using animal tissues. In this study, we employed comparative label-free proteomic analysis for evaluating the solubilization efficacies of the Azo and SDS surfactants using rice leave proteins. This approach led to identification of 3,365 proteins of which 682 proteins were determined as significantly modulated. Further, according to the subcellular localization prediction in SDS and Azo, proteins localized in the chloroplast were the major organelle accounting for 64% of the total organelle in the SDS sample, while only 37.5% of organelle proteins solubilized in the Azo were predicted to be localized in chloroplast. Taken together, this study validates the efficient solubilization of total protein isolated from plant material for bottom-up proteomics. Azo surfactant is suitable as substitute of SDS and promising for bottom-up proteomics as it facilitates robust protein extraction, rapid washing step during enzymatic digestion, and MS analysis.

키워드

과제정보

본 연구는 한국연구재단 중견연구자지원사업 (2019R1A2C2085868) 및 학문후속세대지원사업 (2020R1A6A3A01100427)의 지원으로 수행되었다.

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