생명과학회지 (Journal of Life Science)

  • 제14권2호
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  • Pages.309-314
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  • 2004
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  • 1225-9918(pISSN)
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  • 2287-3406(eISSN)
한국생명과학회 (Korean Society of Life Science)
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온도 기울기 전기영동장치의 CAMP 수용성 단백질에 응용

Application of Temperature Gradient Gel Electrophoresis To cAMP Receptor Protein

  • 발행 : 2004.04.01
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초록

cAMP수용성 단백질(CRP)은 E. coli의 100가지 이상의 유전자 전자조절에 관계된다. CRP는 dimer로 존재하며 cAMP의 결합으로 활성인 형태로 전환된다. 이중체인 CRP 단백질의 열 안정성과 구조 전이의 연구에 효과적인 온도 기울기 전기영동장치를 이용하여 확인하였다. 본 연구에서 야생형과 S83C CRP 단백질의 melting temperature (Tm)는 산성인 완충용액[89.8 mM Glycine, 24mM Boric acid (pH 5.8)]에서 57$\pm$1(야생형 CRP)과 55$\pm$1$^{\circ}C$ (S83G CRP)였다. 그리고 온도에 따른 CRP 단백질의 구조변화도 protease digestion과 CD spectropolarimeter을 이용하여 확인하였다.

Cyclic AMP receptor protein (CRP) is involved in the transcriptional regulation of more than 100 genes in E. coli. CRP dimer is converted into active form via the sequential conformation change of cAMP binding pocket, hinge region and HTH DNA binding motif by binding of cAMP. The temperature gradient gel electrophoresis (TGGE) was applied to CRP protein to know whether it was an efficient technique to study the conformational transitions and the thermal stability. TGGE showed the unfolding process of wild-type and S83G CRP proteins with the temperature gradient set from 29 to 71$^{\circ}C$ on nondenaturing polyacrylamide gel. Melting temperature (Tm) was 57$\pm$1 and 55$\pm$1$^{\circ}C$ for wild-type and S83G CRP, respectively in acidic buffer[89.8 mM Glycine and 24 mM Boric acid (pH 5.8)].

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참고문헌

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