Ordered water molecules as key allosteric mediators in a cooperative dimeric hemoglobin.
Ordered water molecules as key allosteric mediators in a cooperative dimeric hemoglobin.
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DOI：
10.1073/pnas.93.25.14526
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发表时间：
1996-12
影响因子：
11.1
通讯作者：
William E. Royer;A. Pardanani;Quentin H. Gibson;Quentin H. Gibson;Eric S. Peterson;Joel M. Friedman
中科院分区：
文献类型：
--
作者：
William E. Royer;A. Pardanani;Quentin H. Gibson;Quentin H. Gibson;Eric S. Peterson;Joel M. Friedman
一个最显着的结构方面的Scapharca二聚血红蛋白是一个非常有序的水簇在配体结合后，在亚基界面的破坏。我们已经探索了这些晶体学观察到的水分子的作用，通过定点诱变和渗透胁迫技术。界面中Thr-72-->瓦尔的电子等排突变使氧亲和力增加了40倍以上，协同性令人惊讶地增强。这种突变的唯一显著结构效应是使脱氧界面中的两个有序水分子不稳定。野生型Scapharca血红蛋白对渗透压条件非常敏感。加入甘油后，观察到脱氧形式的拉曼光谱的显著变化，表明向配体形式的转变。增加的渗透压，降低了人血红蛋白的氧亲和力，提高了Scapharca血红蛋白的氧亲和力，无论溶质是甘油，葡萄糖还是蔗糖。这些结果的分析提供了一个估计的六个水分子失去后，氧结合的二聚体，在良好的协议，从晶体结构预测的八个。这些实验表明，所观察到的界面水分子簇在亚基之间的通信中起着至关重要的作用。
One of the most remarkable structural aspects of Scapharca dimeric hemoglobin is the disruption of a very well-ordered water cluster at the subunit interface upon ligand binding. We have explored the role of these crystallographically observed water molecules by site-directed mutagenesis and osmotic stress techniques. The isosteric mutation of Thr-72-->Val in the interface increases oxygen affinity more than 40-fold with a surprising enhancement of cooperativity. The only significant structural effect of this mutation is to destabilize two ordered water molecules in the deoxy interface. Wild-type Scapharca hemoglobin is strongly sensitive to osmotic conditions. Upon addition of glycerol, striking changes in Raman spectrum of the deoxy form are observed that indicate a transition toward the liganded form. Increased osmotic pressure, which lowers the oxygen affinity in human hemoglobin, raises the oxygen affinity of Scapharca hemoglobin regardless of whether the solute is glycerol, glucose, or sucrose. Analysis of these results provides an estimate of six water molecules lost upon oxygen binding to the dimer, in good agreement with eight predicted from crystal structures. These experiments suggest that the observed cluster of interfacial water molecules plays a crucial role in communication between subunits.