PDF(Pubmed)
Abstract：
Red blood cells (RBCs) constitute ∼50% of the bloodstream and represent an important target for environmental pollutants and bacterial/viral infections, which can result in their rupture. In addition, diseases such as sickle cell anaemia and paroxysmal nocturnal haemoglobinuria can also result in the rupture of RBCs, which can be potentially life-threatening. With regard to the release of cytosolic metalloproteins from RBCs into the blood-organ system, the biochemical fate of haemoglobin is rather well understood, while comparatively little is known about another highly abundant Zn-metalloprotein, carbonic anhydrase (CA I). To gain insight into the interaction of CA I with human blood plasma constituents, we have employed a metallomics tool comprised of size-exclusion chromatography (SEC) coupled online with an inductively coupled plasma atomic emission spectrometer (ICP-AES), which allows to simultaneously observe all Cu, Fe, and Zn-metalloproteins. After the addition of CA I to human blood plasma incubated at 37°C, the SEC-ICP-AES analysis using phosphate buffered saline (pH 7.4) after 5 min, 1 h, and 2 h revealed that CA I eluted after all endogenous Zn-metalloproteins in the 30 kDa range. Matrix-assisted laser desorption-time of flight mass spectrometry analysis of the collected Zn-peak confirmed that CA I eluted from the column intact. Our in vitro results suggest that CA I released from RBCs to plasma remains free and may be actively involved in health-relevant adverse processes that unfold at the bloodstream-endothelial interface, including atherosclerosis and vision loss.
摘要：
红细胞(RBC)占血液的50%，是环境污染物和细菌/病毒感染的重要目标，这可能会导致它们的破裂。此外,疾病,如镰状细胞性贫血和阵发性夜间血红蛋白尿也可导致红细胞破裂，这可能会危及生命。关于细胞溶质金属蛋白从红细胞释放到血液器官系统中，血红蛋白的生化命运是相当清楚的，虽然对另一种高丰度的锌金属蛋白知之甚少，碳酸酐酶（CAI）。为了深入了解CAI与人血浆成分的相互作用，我们采用了一种金属组学工具，该工具包括与电感耦合等离子体原子发射光谱仪（ICP-AES）在线耦合的尺寸排阻色谱（SEC），可以同时观察所有Cu，铁和锌金属蛋白。在37°C孵育的人血浆中加入CAI后，5分钟后使用磷酸盐缓冲盐水（pH7.4）进行SEC-ICP-AES分析，1小时和2小时显示，CAI在30kDa范围内的所有内源性Zn金属蛋白之后洗脱。收集的Zn峰的基质辅助激光解吸飞行时间质谱分析证实CAI从柱中完整洗脱。我们的体外结果表明，从红细胞释放到血浆的CAI仍然是游离的，并且可能积极参与在血流-内皮界面展开的健康相关不良过程。包括动脉粥样硬化和视力丧失。
