본문 바로가기
HOME> 논문 > 논문 검색상세

논문 상세정보

BMB reports v.43 no.6, 2010년, pp.407 - 412   SCIE
본 등재정보는 저널의 등재정보를 참고하여 보여주는 베타서비스로 정확한 논문의 등재여부는 등재기관에 확인하시기 바랍니다.

Kinetic analysis of Drosophila Vnd protein containing homeodomain with its target sequence

Yoo, Si-Uk    (Department of Biology, College of Sciences, Yeungnam University  );
  • 초록

    Homeodomain (HD) is a highly conserved DNA-binding domain composed of helix-turn-helix motif. Drosophila Vnd (Ventral nervous system defective) containing HD acts as a regulator to either enhance or suppress gene expression upon binding to its target sequence. In this study, kinetic analysis of Vnd binding to DNA was performed. The result demonstrates that DNA-binding affinity of the recombinant protein containing HD and NK2-specific domain (NK2-SD) was higher than that of the full-length Vnd. To access whether phosphorylation sites within HD and NK2-SD affect the interaction of the protein with the target sequence, alanine substitutions were introduced. The result shows that S631A mutation within NK2-SD does not contribute significantly to the DNA-binding affinity. However, S571A and T600A mutations within HD showed lower affinity for DNA binding. In addition, DNA-binding analysis using embryonic nuclear protein also demonstrates that Vnd interacts with other nuclear proteins, suggesting the existence of Vnd as a complex.


  • 주제어

    Analyte recovery .   Drosophila melanogaster .   Homeodomain .   Kinetic analysis .   Vnd.  

  • 참고문헌 (23)

    1. Laughon, A. (1991) DNA Binding Specificity of HDs. Biochemistry 30, 11357-11367. 
    2. Jimenez, F., Martin-Morris, L. E., Velasco, L., Chu, H., Sierra, J., Rosen, D. R. and White, K. (1995) vnd, a gene required for early neurogenesis of Drosophila, encodes a HD protein. EMBO J. 14, 3487-3495. 
    3. Kim, Y. and Nirenberg, M. (1989) Drosophila NK-homeobox genes. Proc. Natl. Acid. Sci. U.S.A. 86, 7716-7720. 
    4. Yu, Z., Syu, L. J. and Mellerick, D. M. (2005) Contextual interactions determine whether the Drosophila HD protein, Vnd, acts as a repressor or activator. Nucleic. Acids. Res. 33, 1-12. 
    5. Gruschus, J. M., Tsao, D. H. H., Wang, L. H., Nirenberg, M. and Ferretti, J. A. (1999) The three-dimensional structure of the vnd/NK-2 HD-DNA complex by NMR spectroscopy. J. Mol. Biol. 289, 529-545. 
    6. Wang, L. H., Chmelik, R. and Nirenberg, M. (2002) Sequence-specific DNA binding by the vnd/NK-2 HD of Drosophila. Proc. Natl. Acid. Sci. U.S.A. 99, 12721-12726. 
    7. Stepchenko, A. and Nirenberg, M. (2004) Mapping activation and repression domains of the vnd/NK-2 HD protein. Proc. Natl. Acid. Sci. U.S.A. 101, 13180-13185. 
    8. Nirenberg, M., Nakayama, K., Nakayama, N., Kim, Y., Mellerick, D., Wang, L. H., Webber, K. O. and Lad, R. (1995) The NK-2 homeobox gene and the early development of the central nervous system of Drosophila. Ann. N.Y. Acad. Sci. 758, 224-242. 
    9. Gruschus, J. M., Tsao, D. H. H., Wang, L. H., Nirenberg, M. and Ferretti, J. A. (1997) Interactions of the vnd/NK-2 HD with DNA by nuclear magnetic resonance spectroscopy: basic of binding specificity. Biochemistry 36, 5372-5380. 
    10. Weiler, S., Gruschus, J. M., Tsao, D. H. H., Yu, L., Wang, L. H., Nirenberg, M. and Ferretti, J. A. (1998) Site-directed mutations in the vnd/NK-2 HD. J. Biol. Chem. 273, 10994-11000. 
    11. Karlsson, R. and Falt, A. (1997) Experimental design for kinetic analysis of protein-protein interactions with surface plasmon resonance biosensors. J. Immunological Methods 200, 121-133. 
    12. Katsamba, P. S., Park, S. and Laird-Offringa I. A., (2002) Kinetic studies of RNA-protein interactions using surface plasmon resonance. Methods 26, 95-104. 
    13. Pan, D. J., Huang, J. D. and Courey, A. J. (1991) Functional analysis of the Drosophila twist promoter reveals a dorsal-binding ventral activator region. Genes Dev. 10, 1892-1901. 
    14. Park, S., Myszka, D. G., Yu, M., Littler, S. J. and Laird-Offringa, I. A. (2000) HuD RNA recognition motifs play distinct roles in the formation of a stable complex with AU-rich RNA. Mol. Cell. Biol. 20, 4765-4772. 
    15. Tsao, D. H. H., Gruschus, J. M., Wang, L. H., Nirenberg, M. and Ferretti, J. A. (1994) Elongation of helix III of the NK-2 HD upon binding to DNA: a secondary structure study by NMR. Biochemistry 33, 15053-15060. 
    16. Koizumi, K., Lintas, C., Nirenberg, M., Maeng, J. S., Ju, J. H., Mack, J. W., Gruschus, J. M., Odenwald, W. F. and Ferretti, J. A. (2003) Mutations that affect the ability of the vnd/NK-2 HD to regulate gene expression: transgenic alterations and tertiary structure. Proc. Natl. Acid. Sci. U.S.A. 100, 3119-3124. 
    17. Zhang, H., Syu, L. J., Modica, V., Yu, Z., Ohlen, T. V. and Mellerick, D. M. (2008) The Drosophila HD transcription factor, Vnd, associates with a variety of co-factors, is extensively phosphorylated and forms multiple complexes in embryos. FEBS J. 275, 5062-5073. 
    18. Uhler, J., Zhang, H, Syu, L. J. and Mellerick, D. M. (2007) The Nk-2 box of the Drosophila HD protein, Vnd, contributes to its repression activity in a Groucho-dependent manner. Mech. Dev. 124, 1-10. 
    19. Watada, H., Mirmira, R. G., Kalamaras, J. and German, M. S. (2000) Intramolecular control of transcriptional activity by the NK-2 specific domain in NK-2 HD proteins. Proc. Natl. Acid. Sci. U.S.A. 97, 9443-9448. 
    20. Sonksen, C. P., Nordhoff, E., Jansson, O., Malmqvist, M. and Roepstorff, P. (1998) Combining MALDI mass spectrometry and biomolecular interaction analysis using a biomolecular interaction analysis instrument. Anal. Chem. 70, 2731-2736. 
    21. Zhukov, A., Schurenberg, M., Jansson, O., Areskoug, D. and Buijs, J. (2004) Integration of surface plasmon resonance with mass spectrometry: automated ligand fishing and sample preparation for MALDI MS using a Biacore 3000 biosensor. J. Biomol. Tech. 15, 112-119. 
    22. Vermette, P., Gengenbach, T., Divisekera, U., Kambouris, P. A., Griesser, H. J. and Meagher, L. (2003) Immobilization and surface characterization of NeutrAvidin biotin-binding protein on different hydrogel interlayers. J. Colloid Interface Sci. 259, 13-26. 
    23. Fisher, P. A. and Blobel, G. (1983) Preparation of a nuclear matrix-pore complex-lamina fraction from embryos of Drosophila melanogaster. Methods Enzymol. 96, 589-596. 

 저자의 다른 논문

 활용도 분석

  • 상세보기

    amChart 영역
  • 원문보기

    amChart 영역

원문보기

무료다운로드
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