본문 바로가기
HOME> 논문 > 논문 검색상세

논문 상세정보

The journal of microbiology v.43 no.3, 2005년, pp.295 - 300   피인용횟수: 3

Comparative Evaluation of Three Purification Methods for the Nucleocapsid Protein of Newcastle Disease Virus from Escherichia coli Homogenates

Tan Yan Peng    (Department of Chemical and Environmental Engineering, Institute of Bioscience, University Putra Malaysia   ); Ling Tau Chuan    (Department of Process & Food Engineering, Faculty of Engineering, Institute of Bioscience, University Putra Malaysia   ); Yusoff Khatijah    (Institute of Bioscience, Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, University Putra Malaysia   ); Tan Wen Siang    (Institute of Bioscience, Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, University Putra Malaysia   ); Tey Beng Ti    (Department of Chemical and Environmental Engineering, Institute of Bioscience, University Putra Malaysia  );
  • 초록

    In the present study, the performances of conventional purification methods, packed bed adsorption (PBA), and expanded bed adsorption (EBA) for the purification of the nucleocapsid protein (NP) of Newcastle disease virus (NDV) from Escherichia coli homogenates were evaluated. The conventional methods for the recovery of NP proteins involved multiple steps, such as centrifugation, precipitation, dialysis, and sucrose gradient ultracentrifugation. For the PBA, clarified feedstock was used for column loading, while in EBA, unclarified feedstock was used. Streamline chelating immobilized with $Ni^{2+}$ ion was used as an affinity ligand for both PBA and EBA. The final protein yield obtained in conventional and PBA methods was $1.26\%$ and $5.56\%$ , respectively. It was demonstrated that EBA achieved the highest final protein yield of $9.6\%$ with a purification factor of 7. Additionally, the total processing time of the EBA process has been shortened by 8 times compared to that of the conventional method.


  • 주제어

    NP protein .   NDV .   Escherichia coli .   expanded bed adsorption .   packed bed adsorption.  

  • 참고문헌 (25)

    1. Ameskamp, N., C. Priesner, J. Lehmann, and D. Lutkemeyer. 1999. Pilot scale recovery of monoclonal antibodies by expanded bed ion exchange adsorption. Bioseparation 8, 169-88 
    2. Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248-254 
    3. Laemmli, U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680-685 
    4. Noppe, W., I. Hanssens, and M.D. Cuyper. 1996. Simple two step procedure for the preparation of highly active pure equine milk lysozyme. J. Chromatogr. A 719, 327-331 
    5. Ueda, E.K.M., P.W. Gout, and L. Morganti. 2003. Current and prospective applications of metal ion-protein binding. J. Chromatogr. A 988, 1-23 
    6. Thommes, J., A. Bader, M. Halfar, A. Karau, and M-R Kula. 1996. Isolation of monoclonal antibodies from cell containing hybridoma broth using a protein A. coated adsorbent in expanded beds. J. Chromatogr. A 752, 111-122 
    7. Batt, B.C., V.M. Yabannavar, and V. Singh. 1995. Expanded bed adsorption process for protein recovery from whole mammalian cell culture broth. Bioseparation 5, 41-52 
    8. Chaga, G.S. 2001. Twenty-five years of immobilized metal ion affinity chromatography: past, present and future. J. Biochem. Biophys. Methods 49, 313-334 
    9. Kaufmann, M. 1997. Unstable proteins: how to subject them to chromatographic separations for purification procedures. J. Chromatogr. B 699, 347-369 
    10. Yusoff, K. and W.S. Tan. 2001. Newcastle disease virus: macromolecules and opportunities. Avian Pathol. 30, 439-455 
    11. Hu, Y.C., C. T. Tsai, Y.C. Chung, J.T. Lu, and J.T. Hsu. 2003. Generation of chimeric baculovirus with histidine-tags displayed on the envelope and its purification using immobilized metal affinity chromatography. Enzyme Microb. Technol. 33, 445-452 
    12. Chase, H.A. and N.M. Draeger. 1992. Affinity purification of proteins using expanded beds. J. Chromatogr. A 597,129-145 
    13. Ling, T.C., C.K. Long, W.S. Tan, B.T. Tey, A. Ariff, W.M. Wan Abdullah. 2004. Purification of filamentous bacteriophage M13 by expanded bed anion exchange chromatography. J. Micorbiol. 42, 22-232 
    14. Xiang, H., R. Wynn, L-H. T. Nguyen, O.H. Ross, D.P. Ahrens, K.T. O'Neil, G.F. Hollis, and D.R. Patrick. 2002. Immobilized metal-ion affinity chromatography of recombinant Fab protein OPG C11 in the presence of EDTA-Mg (II). J. Chromatogr. A 978, 153-164 
    15. Lee, S.M. 1989. The primary stages of protein recovery. J. Biotechnol. 11, 103-118 
    16. Chang, Y.K. and H.A. Chase. 1996. Ion exchange purification of G6PDH from unclarified yeast cell homogenates using expanded bed adsorption. Biotechnol. Bioeng. 49, 204-216 
    17. Ersson, B., L. Ryden, and J.-C. Janson. 1998. Introduction to protein purification, p. 3 - 40. In J-C. Janson, L. Ryden (eds.), Protein Purification: Principles, High-Resolution Methods, and Applications. John Wiley & Sons, New York 
    18. Ling, T.C. 2002. Ph. D. thesis. The University of Birmingham, UK 
    19. Rabu, A., W.S. Tan, C.L. Kho, A.R. Omar, and K. Yusoff. 2002. Chimeric Newcastle disease virus nucleocapsid with parts of its viral haemagglutinin-neuraminidase and fusion proteins. Acta Virol. 46, 211-217 
    20. Willoughby, N.A., T. Kirschner, M.P. Smith, R. Hjorth, and N.J. Titchener-Hooker. 1999. Immobilized metal affinity chromatography purification of alcohol dehydrogenase from Baker's yeast using an expanded bed adsorption system. J. Chromatogr. A 840, 195-204 
    21. Kho, C.L., W.S. Tan, and K. Yusoff. 2001. Production of the nucleocapsid protein of Newcastle disease virus in Escherichia coli and its assembly into ring- and nucleocapsid-like particles. J. Microbiol. 39, 293-299 
    22. Thommes, J., M. Halfar, S. Lenz, M.-R. Kula. 1995. Purification of monoclonal antibodies from whole hybridoma fermentation broth by fluidized bed adsorption. Biotechnol. Bioeng. 45, 205-211 
    23. Anspach, F.B., D. Curbelo, R. Hartmann, G. Garke, and W.D. Deekwer. 1999. Expanded-bed chromatography in primary protein purification. J. Chromatogr. A 865, 129-144 
    24. Tey, B.T., K.H.Yong, H.P. Ong, T.C. Ling, S.T. Ong, Y.P. Tan, A. Ariff, and W.S. Tan. 2004. Optimal conditions for Hepatitis B core Antigen production in shaked flask fermentation. Biotechnol Bioprocess Eng. 9, 374-378 
    25. Sumi, A., K. Okuyama, K. Kobayashi, W. Ohtani, T. Ohmura, and K. Yokoyama. 1999. Purification of recombinant human serum albumin efficient purification using STREAMLINE. Bioseparation 8, 195-200 
  • 이 논문을 인용한 문헌 (3)

    1. 2006. "" Biotechnology and bioprocess engineering, 11(2): 164~167     
    2. 2006. "" Biotechnology and bioprocess engineering, 11(3): 268~272     
    3. 2009. "" 大韓獸醫學會誌 = Korean journal of veterinary research, 49(1): 39~44     

 활용도 분석

  • 상세보기

    amChart 영역
  • 원문보기

    amChart 영역

원문보기

무료다운로드
  • NDSL :
유료다운로드

유료 다운로드의 경우 해당 사이트의 정책에 따라 신규 회원가입, 로그인, 유료 구매 등이 필요할 수 있습니다. 해당 사이트에서 발생하는 귀하의 모든 정보활동은 NDSL의 서비스 정책과 무관합니다.

원문복사신청을 하시면, 일부 해외 인쇄학술지의 경우 외국학술지지원센터(FRIC)에서
무료 원문복사 서비스를 제공합니다.

NDSL에서는 해당 원문을 복사서비스하고 있습니다. 위의 원문복사신청 또는 장바구니 담기를 통하여 원문복사서비스 이용이 가능합니다.

이 논문과 함께 출판된 논문 + 더보기