The role of phenylalanine 31 in maintaining the conformational stability of ribonuclease P2 from Sulfolobus solfataricus under extreme conditions of temperature and pressure
The role of phenylalanine 31 in maintaining the conformational stability of ribonuclease P2 from Sulfolobus solfataricus under extreme conditions of temperature and pressure
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DOI：
10.1021/bi970467v
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发表时间：
1997-07-22
期刊：
影响因子：
2.9
通讯作者：
Lange, R
中科院分区：
文献类型：
--
作者：
Mombelli, E;Afshar, M;Lange, R
来自嗜热古细菌Sulfolobus solfataricus的核糖核酸酶P2是一种已知三维结构的小蛋白（7kda）。结构检测和分子动力学模拟表明，疏水核中的3个芳香残基（Phe5、Phe31和Tyr33）具有较强的范德华相互作用能。通过分析四阶导数模式下蛋白质的紫外吸收，研究了野生型和F31A、F31Y突变体在热、冷、压力诱导下蛋白质构象变化的热力学。野生型蛋白在各种温度和压力条件下都非常稳定。两个突变体的热变性和冷变性，以及F31A突变体的压力变性，导致导数光谱的显著蓝移，表明Tyr33的溶剂暴露增加。对于F31Y突变体，高压（400 MPa）保护蛋白质免受热变性。这项研究，探索疏水芳香核心的性质，补充了热展开研究，探索整体结构变化[Knapp， S., Karshikoff, a ., Berndt， K. D., Christova， P., Atanasov， B., & Ladenstein， R. (1996) J. Mol. Biol. 264, 1132-1144]。在极端温度、压力和pH下观察到的差异可能是一种展开机制的合理解释，该机制涉及外周蛋白的大部分，同时保持疏水核心的完整性。
Ribonuclease P2 from the thermophilic archaebacterium Sulfolobus solfataricus is a small protein (7 kDa) with a known three-dimensional structure. Inspection of the structure and molecular dynamics simulation reveal that three aromatic residues (Phe5, Phe31, and Tyr33) from the hydrophobic core have a strong van der Waals interaction energy. We studied the thermodynamics of the heat, cold, and pressure-induced protein conformational changes of the wild type and of the F31A and F31Y mutants by analyzing the protein UV absorbance in the fourth derivative mode. The wild-type protein was extremely stable under all conditions of temperature and pressure. Heat and cold denaturation of both mutants, as well as denaturation by pressure of the F31A mutant, led to significant blue shifts of the derivative spectrum, indicating increased solvent exposure of Tyr33. For the F31Y mutant, high pressure (400 MPa) protected the protein against thermal denaturation. This study, probing the properties of the hydrophobic aromatic core, complements a thermal unfolding study which probes the overall structural changes [Knapp, S., Karshikoff, A., Berndt, K. D., Christova, P., Atanasov, B., & Ladenstein, R. (1996) J. Mol. Biol. 264, 1132-1144]. The differences observed in response to extremes of temperature, pressure, and pH may be rationalized by an unfolding mechanism involving larger parts of the peripheral protein while the integrity of the hydrophobic core is maintained.