RESPONSE OF A PROTEIN-STRUCTURE TO CAVITY-CREATING MUTATIONS AND ITS RELATION TO THE HYDROPHOBIC EFFECT
RESPONSE OF A PROTEIN-STRUCTURE TO CAVITY-CREATING MUTATIONS AND ITS RELATION TO THE HYDROPHOBIC EFFECT
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DOI：
10.1126/science.1553543
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发表时间：
1992-01-10
期刊：
影响因子：
56.9
通讯作者：
MATTHEWS, BW
中科院分区：
文献类型：
--
作者：
ERIKSSON, AE;BAASE, WA;MATTHEWS, BW
在噬菌体T4溶菌酶的疏水核心内构建了6种“产生空腔”的突变体，即亮氨酸46→丙氨酸（L46A）、L99A、L118A、L121A、L133A以及苯丙氨酸153→丙氨酸（F153A）。这些取代使蛋白质在pH 3.0时的稳定性降低的程度不同，L46A和L121A降低了2.7千卡/摩尔（kcal mol⁻¹），而L99A降低了5.0 kcal mol⁻¹。还构建了双突变体L99A/F153A，其稳定性降低了8.3 kcal mol⁻¹。所有变体的X射线结构都以高分辨率确定。在每种情况下，野生型侧链的去除使一些周围的原子向空出的空间移动，但总是会留下一个空腔，其体积从L46A的24立方埃（ų）到L99A的150 ų不等。在这些空腔中均未观察到溶剂分子。相对于野生型，突变的亮氨酸→丙氨酸蛋白质的去稳定作用可以用一个常数项（约2.0 kcal mol⁻¹）加上一个与空腔大小成比例增加的项来近似。该常数项大约等于根据水相和有机相之间的分配所确定的亮氨酸相对于丙氨酸的转移自由能。随空腔大小增加的能量项可以用空腔体积（24 - 33 cal mol⁻¹ Å⁻³）或空腔表面积（20 cal mol⁻¹ Å⁻²）来表示。这些结果提示了如何调和关于蛋白质中疏水作用强度的许多相互矛盾的报道。
Six "cavity-creating" mutants, LeU46 --> Ala (L46A), L99A, L118A, L121A, L133A, and Phe153 --> Ala (F153A), were constructed within the hydrophobic core of phage T4 lysozyme. The substitutions decreased the stability of the protein at pH 3.0 by different amounts, ranging from 2.7 kilocalories per mole (kcal mol-1) for L46A and L121A to 5.0 kcal mol-1 for L99A. The double mutant L99A/F153A was also constructed and decreased in stability by 8.3 kcal mol-1. The x-ray structures of all of the variants were determined at high resolution. In every case, removal of the wild-type side chain allowed some of the surrounding atoms to move toward the vacated space but a cavity always remained, which ranged in volume from 24 cubic angstroms (angstrom 3) for LA6A to 150 angstrom 3 for L99A. No solvent molecules were observed in any of these cavities. The destabilization of the mutant Leu --> Ala proteins relative to wild type can be approximated by a constant term (approximately 2.0 kcal mol-1) plus a term that increases in proportion to the size of the cavity. The constant term is approximately equal to the transfer free energy of leucine relative to alanine as determined from partitioning between aqueous and organic solvents. The energy term that increases with the size of the cavity can be expressed either in terms of the cavity volume (24 to 33 cal mol-1 angstrom -3) or in terms of the cavity surface area (20 cal mol-1 angstrom -2). The results suggest how to reconcile a number of conflicting reports concerning the strength of the hydrophobic effect in proteins.