Luka Zigmund, Wagner Conrad
Department of Biochemistry, Medical Center, Vanderbilt University, School of Medicine, 620 Light Hall, Nashville, TN 37232-0146, USA.
Arch Biochem Biophys. 2003 Dec 1;420(1):153-60. doi: 10.1016/j.abb.2003.09.009.
Human recombinant glycine N-methyltransferase (GNMT) unfolding by urea was studied by enzyme activity, size-exclusion chromatography, fluorescence spectroscopy, and circular dichroism. Urea unfolding of GNMT is a two-step process. The first transition is a reversible dissociation of the GNMT tetramer to compact monomers in 1.0-3.5M urea with enzyme inactivation. The compact monomers were characterized by Stokes radius (R(s)) of 40.7A equal to that of globular proteins with the same molecular mass as GNMT monomers, absence of exposure of tryptophan residues into solvent, and presence of about 50% of secondary structure of native protein. The second step of GNMT unfolding is a reversible transition of monomers from compact to a fully unfolded state with R(s) of 50A, exposed tryptophan residues, and disrupted secondary structure in 8M urea.
通过酶活性、尺寸排阻色谱、荧光光谱和圆二色性研究了尿素诱导的人重组甘氨酸N-甲基转移酶（GNMT）的去折叠过程。GNMT的尿素去折叠是一个两步过程。第一步转变是在1.0 - 3.5M尿素中，GNMT四聚体可逆解离为紧密单体，同时酶失活。紧密单体的斯托克斯半径（R(s)）为40.7Å，与具有和GNMT单体相同分子量的球状蛋白的斯托克斯半径相等，色氨酸残基未暴露于溶剂中，且具有约50%的天然蛋白二级结构。GNMT去折叠的第二步是单体从紧密状态可逆转变为完全去折叠状态，此时R(s)为50Å，色氨酸残基暴露，二级结构在8M尿素中被破坏。
