지원사업
학술연구/단체지원/교육 등 연구자 활동을 지속하도록 DBpia가 지원하고 있어요.
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연구자들이 자신의 연구와 전문성을 널리 알리고, 새로운 협력의 기회를 만들 수 있는 네트워킹 공간이에요.
논문 기본 정보
- 자료유형
- 학술저널
- 저자정보
- 발행연도
- 2013.7
- 수록면
- 39 - 46 (8page)
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초록· 키워드
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Proteins have evolved to compensate for detrimental mu-tations. However, compensatory mechanisms for protein defects are not well understood. Using ketosteroid isome-rase (KSI), we investigated how second-site mutations could recover defective mutant function and stability. Pre-vious results revealed that the Y30F mutation rescued the Y14F, Y55F and Y14F/Y55F mutants by increasing the catalytic activity by 23-, 3- and 1.3-fold, respectively, and the Y55F mutant by increasing the stability by 3.3 kcal/mol. To better understand these observations, we systematically investigated detailed structural and thermodynamic effects of the Y30F mutation on these mutants. Crystal structures of the Y14F/Y30F and Y14F/Y55F mutants were solved at 2.0 and 1.8 A resolution, respectively, and compared with previoulsy solved structures of wild-type and other mutant KSIs. Structural analyses revealed that the Y30F mutation partially restored the active-site cleft of these mutant KSIs. The Y30F mutation also increased Y14F and Y14F/Y55F mutant stability by 3.2 and 4.3 kcal/mol, respectively, and the melting temperatures of the Y14F, Y55F and Y14F/Y55F mutants by 6.4°C, 5.1°C and 10.0°C, respectively. Compensatory effects of the Y30F mutation on stability might be due to improved hydrophobic interactions because removal of a hydroxyl group from Tyr30 induced local compaction by neighboring residue movement and enhanced interactions with surrounding hydrophobic residues in the active site. Taken together, our results suggest that perturbed active-site geometry recovery and favorable hydrophobic interactions mediate the role of Y30F as a second-site suppressor.
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참고문헌 (32)
참고문헌 신청
Blance, S.J / 2000 / Temperature-sensitive suppressor mutations of the Escherichia coli DNA gyrase B protein / Protein Sci 9:1035~1037
Brachmann, R.K / 1998 / Genetic selection of intragenic suppressor mutations that reverse the effect of common p53 cancer mutations / EMBO J 17:1847~1859
Brunger, A.T / 1998 / Crystallography & NMR system: a new software suite for macromolecular structure determination / Acta Crystallogr. D Biol. Crystallogr 54:905~921
Cho, H.S / 1999 / Crystal structure of delta(5)-3-ketosteroid isomerase from Pseudomonas testosteroni in complex with equilenin settles the correct hydrogen bonding scheme for transition state stabilization / J. Biol. Chem 274:32863~32868
Choi, G. / 2000 / Asp-99 donates a hydrogen bond not to Tyr- 14 but to the steroid directly in the catalytic mechanism of Delta 5-3-ketosteroid isomerase from Pseudomonas putida biotype B / Biochemistry 39:903~909
Brachmann, R.K / 1998 / Genetic selection of intragenic suppressor mutations that reverse the effect of common p53 cancer mutations / EMBO J 17:1847~1859
Brunger, A.T / 1998 / Crystallography & NMR system: a new software suite for macromolecular structure determination / Acta Crystallogr. D Biol. Crystallogr 54:905~921
Cho, H.S / 1999 / Crystal structure of delta(5)-3-ketosteroid isomerase from Pseudomonas testosteroni in complex with equilenin settles the correct hydrogen bonding scheme for transition state stabilization / J. Biol. Chem 274:32863~32868
Choi, G. / 2000 / Asp-99 donates a hydrogen bond not to Tyr- 14 but to the steroid directly in the catalytic mechanism of Delta 5-3-ketosteroid isomerase from Pseudomonas putida biotype B / Biochemistry 39:903~909
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