Dybas Jakub, Bokamper Matthew J, Marzec Katarzyna M, Mak Piotr J
Saint Louis University, Chemistry Department, 3501 Laclede Ave., 63103 Saint Louis, MO, United States; Jagiellonian University, Jagiellonian Centre for Experimental Therapeutics (JCET), 14 Bobrzyńskiego Str., 30-348 Krakow, Poland.
Saint Louis University, Chemistry Department, 3501 Laclede Ave., 63103 Saint Louis, MO, United States.
Spectrochim Acta A Mol Biomol Spectrosc. 2020 Oct 5;239:118530. doi: 10.1016/j.saa.2020.118530. Epub 2020 May 29.
Hemoglobin (Hb) is a key component of respiratory system and as such plays important role in human physiology. The studies of Hb's structure and functions are usually performed on cell-free protein; however, it has been shown that there are functionally relevant differences between isolated Hb and Hb present inside red blood cells (RBCs). It is clear that new experimental approaches are needed to understand the origin of these differences and to gain insight into the structure-function relationship of Hb within intact living cells. In this work we present a novel application of Resonance Raman spectroscopy to study heme active site of different forms of human Hb within living RBCs using laser excitation lines in resonance with their Soret absorption bands. These studies revealed that there are no significant changes in the disposition of the Fe-O-O fragment or the Fe-N linkage for Hb molecules enclosed in RBCs and these in free isolated states. However, some changes in the orientation of the heme vinyl groups were observed which might account for the differences in the protein activity and ligand affinity. This work highlights importance of protein-based studies and presents a new opportunity to translate these results to physiological cell systems.
血红蛋白（Hb）是呼吸系统的关键组成部分，因此在人体生理学中发挥着重要作用。对Hb结构和功能的研究通常在无细胞蛋白质上进行；然而，已经表明，分离的Hb与红细胞（RBC）内存在的Hb在功能上存在相关差异。显然，需要新的实验方法来理解这些差异的起源，并深入了解完整活细胞内Hb的结构-功能关系。在这项工作中，我们展示了共振拉曼光谱的一种新应用，即利用与它们的Soret吸收带共振的激光激发线，研究活RBC内不同形式的人Hb的血红素活性位点。这些研究表明，对于包裹在RBC中的Hb分子以及处于游离分离状态的Hb分子，Fe-O-O片段或Fe-N键的排列没有显著变化。然而，观察到血红素乙烯基的方向有一些变化，这可能解释了蛋白质活性和配体亲和力的差异。这项工作突出了基于蛋白质研究的重要性，并为将这些结果转化到生理细胞系统提供了新机会。
