Studies of Pseudomonas aeruginosa azurin mutants: cavities in beta-barrel do not affect refolding speed.
Studies of Pseudomonas aeruginosa azurin mutants: cavities in beta-barrel do not affect refolding speed.
复制标题
铜绿假单胞菌天青蛋白突变体的研究：β-桶中的空腔不影响重折叠速度。
DOI：
10.1016/s0006-3495(02)75606-3
复制
发表时间：
2002
影响因子：
3.4
通讯作者：
Wittung-Stafshede,Pernilla
中科院分区：
文献类型：
--
作者：
Pozdnyakova,Irina;Guidry,Jesse;Wittung-Stafshede,Pernilla
铜绿假单胞菌是一种具有希腊键折叠的蓝铜蛋白。去除铜会产生与全氮蛋白结构相同的载脂蛋白。为了解决β-桶中小空腔对热力学稳定性和折叠动力学的影响，我们研究了野生型和两个突变体（His-46-Gly和His-117-Gly） azurins的载子形式的行为。这三种蛋白质的平衡和动力学折叠和展开反应表现为两态过程。这两个突变体的热力学稳定性与野生型azurin的稳定性相比明显降低，这与疏水内部或附近的空腔具有整体不稳定作用一致。在展开速度上也发现了巨大的差异：突变体比野生型azurin展开得快得多。相比之下，这三种蛋白质的折叠速率常数几乎相同，并且与azurin的原生态拓扑预测的速率常数密切匹配。我们得出结论，在决定azurin折叠速度方面，拓扑结构比平衡稳定性更重要。
Pseudomonas aeruginosaazurin is a blue-copper protein with a Greek-key fold. Removal of copper produces an apoprotein with the same structure as holoazurin. To address the effects on thermodynamic stability and folding dynamics caused by small cavities in aβ-barrel, we have studied the behavior of the apo-forms of wild-type and two mutant (His-46-Gly and His-117-Gly) azurins. The equilibrium- and kinetic-folding and unfolding reactions appear as two-state processes for all three proteins. The thermodynamic stability of the two mutants is significantly decreased as compared with the stability of wild-type azurin, in accord with cavities in or near the hydrophobic interior having an overall destabilizing effect. Large differences are also found in the unfolding rates: the mutants unfold much faster than wild-type azurin. In contrast, the folding-rate constants are almost identical for the three proteins and closely match the rate-constant predicted from the native-state topology of azurin. We conclude that the topology is more important than equilibrium stability in determining the folding speed of azurin.