Division of Physiological Chemistry I, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Scheelesväg 2, SE-17 177 Stockholm, Sweden.
J Am Soc Mass Spectrom. 2011 Oct;22(10):1763-70. doi: 10.1007/s13361-011-0182-0. Epub 2011 Jul 19.
His64 and His93 are the two well-known sites of heme binding in water-dissolved holo-myoglobin, with His93 being a proximal, strongly binding partner, while the distal His64 weakly coordinates to the heme through a small-molecule ligand, e.g., water or O(2). The heme bonding scheme in a water-free environment is as yet unclear. Here we employed electron transfer dissociation tandem mass spectrometry to study the preferential attachment site of the ferri-heme (Fe(3+)) in electrospray-produced 12+, 14+, and 16+ holo-myoglobin ions. Contrary to expectations, in lower-charge complexes that should have a structure resembling that in solution, the heme seems to be preferentially attached to the "distal" histidine. In contrast, in the highest studied charge state, the "proximal" histidine is the site of preferential attachment; the 14+ charge state is an intermediate case. This surprising finding raises a question of heme coordination in proteins transferred to water-free environment, as well as the effect of the protonation sites on heme bonding.
His64 和 His93 是水合血红素-肌红蛋白中两个众所周知的血红素结合位点，His93 是一个紧邻的、强结合配体，而远端的 His64 通过小分子配体（如水或 O(2)）弱配位到血红素上。在无水环境中，血红素的结合方案尚不清楚。在这里，我们采用电子转移解离串联质谱法研究了电喷雾产生的 12+、14+和 16+ 血红素-肌红蛋白离子中 ferri-heme(Fe(3+))的优先附着位点。与预期相反，在电荷较低的配合物中，其结构应类似于溶液中的结构，血红素似乎优先附着在“远端”组氨酸上。相比之下，在研究的最高电荷状态下，“近端”组氨酸是优先附着的位点；14+ 电荷状态是一个中间情况。这一惊人的发现提出了一个问题，即在转移到无水环境的蛋白质中，血红素的配位以及质子化位点对血红素键合的影响。
