游离芳香氨基酸和半胱氨酸对光诱导高铁肌红蛋白还原的影响

肌红蛋白(Myoglobin,Mb)中血红素辅基不仅具有储氧功能,也能吸收特定波长的光而影响蛋白功能表达。实验发现,部分游离的氨基酸对光诱导高铁肌红蛋白(metM b,Fe(III)-Mb)的还原过程及还原程度都有重要作用,因此,本文采用紫外-可见吸收光谱、圆二色谱、三维荧光光谱法,在光照体系中加入拥挤试剂来模拟细胞内拥挤环境,研究芳香氨基酸色氨酸(Trp)、苯丙氨酸(Phe)、酪氨酸(Tyr)、半胱氨酸(Cys)]对metM b还原的影响。结果表明,含-OH或-SH的氨基酸(Tyr、Cys)能使metM b发生较好的光还原,无-OH或-SH基团的氨基酸(Trp、Phe)对metM b的光还原作用较弱,氨基酸促进metM b光还原的整个过程可能是分子间电子转移的过程。metM b在拥挤环境聚蔗糖70(Ficoll 70)中的光诱导还原程度比在稀溶液中高,拥挤试剂Ficoll70对蛋白的二级结构起保护作用,能够稳定血红素微环境。

Effect of Free Aromatic Amino Acids and Cysteine on Photoreduction of met-Myoglobin

Myoglobin heme cofactor can absorb specific wavelength of light to affect the expression of protein function. In this work, we found free amino acid played an important role in photoreduction of met-myoglobin. Applied with UV-Vis absorption, CD spectra and three-dimensional fluorescence spectroscopy, we studied the aromatic amino acids (tryptophan, phenylalanine and tyrosine) and cysteine impacts on the photoreduction of myoglobin in the crowded environment by adding crowded reagents. The UV-Vis absorption spectrum data revealed that the amino acids containing -OH or -SH groups (Tyr, Cys) could facilitate the photoreduction process of Mb, while the Amino acids (Trp, Phe) without -OH or -SH groups had weakly effect on photoreduction of Mb. The amino acid promotion of the Mb photoreduction process may be radical electron transfer process. The photoreduction extent of metMb was much higher in Ficoll70 solution than that in dilute solution. In addition, crowded reagent Ficoll70 could protect the secondary structure of protein and stabilize the heme microenvironment.