Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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The β Subunit of Heterotrimeric G Protein Interacts Directly with Kinesin Heavy Chains, Kinesin-I

Kinesin-I의 kinesin heavy chains과 직접 결합하는 heterotrimeric G protein의 β subunit의 규명

  • Seog, Dae-Hyun (Departments of Biochemistry, College of Medicine, Inje University)
  • 석대현 (인제대학교 의과대학 생화학교실)
  • Received : 2010.05.19
  • Accepted : 2010.06.14
  • Published : 2010.08.30
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Abstract

Kinesin-I exists as a tetramer of two heavy chains (KHCs, also called KIF5s), which contain the amino (N)-terminal motor domain and carboxyl (C)-terminal domain, as well as two light chains (KLCs), which bind to the KIF5s (KIF5A, KIF5B and KIF5C) stalk region. To identify the interaction proteins for KIF5A, yeast two-hybrid screening was performed and a specific interaction with the ${\beta}$ subunit of heterotrimeric G proteins ($G{\beta}$) was found. $G{\beta}$ bound to the amino acid residues between 808 and 935 of KIF5A and to other KIF5 members in the yeast two-hybrid assay. The WD40 repeat motif of $G{\beta}$ was essential for interaction with KIF5A. In addition, these proteins showed specific interactions in the glutathione S-transferase (GST) pull-down assay. An antibody to KIF5s specifically co-immunoprecipitated KIF5s associated with heterotrimeric G proteins from mouse brain extracts. These results suggest that kinesin-I motor protein transports heteroterimeric G protein attachment vesicles along microtubules in the cell.

Kinesin-I은 4분자의 단백질로 구성되어 있으며, N-말단의 motor 영역과 C-말단영역을 가지는 장쇄(KHC, 또한 KIF5s로도 통용) 2분자와 KIF5s (KIF5A, KIF5B와 KIF5C)의 줄기영역과 결합하는 단쇄(KLC) 2분자로 구성되어 있다. KIF5A의 결합 단백질을 동정하기 위하여 효모 two-hybrid system을 사용하여 특이적으로 결합하는 heterotrimeric G 단백질의 ${\beta}$ 단위체 단백질($G{\beta}$)을 분리하였다. $G{\beta}$은 KIF5A의 808에서 935아미노산 부위와 결합하며, 다른 KIF5들과도 결합함을 효모 two-hybrid assay로 확인하였다. 또한 $G{\beta}$의 WD40 반복 서열은 KIF5A와의 결합에 필수영역임을 확인하였으며, 이러한 단백질간의 결합은 Glutathione S-transferase (GST) pull-down assay를 통하여 확인하였다. 생쥐의 뇌 파쇄액에 KIF5들의 항체로 면역침강을 행하여 heterotrimeric G 단백질을 확인한 결과, KIF5들은 heterotrimeric G 단백질과 특이적으로 같이 침강하였다. 이러한 결과들은 kinesin-I는 heterotrimeric G 단백질이 포함된 소포를 미세소관을 따라 이동시킴을 시사한다.

Keywords

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