Food Research Institute, National Agriculture and Food Research Organization, 2-1-12 Kannondai, Tsukuba, Ibaraki, 305-8642, Japan.
J Chromatogr A. 2019 Oct 11;1603:190-198. doi: 10.1016/j.chroma.2019.06.042. Epub 2019 Jun 22.
The heat-induced (80 ± 1 ℃, 1 h) aggregates of bovine serum albumin and ovalbumin at neutral pH and low ionic strength (10 mM sodium phosphate buffer, pH 6.9) were characterized using size-exclusion chromatography combined with small-angle X-ray scattering measurement. The values calculated for the radius of gyration and molecular weight of the eluted aggregates of the bovine serum albumin were 9-11 nm and 540,000-820,000, respectively. Those of ovalbumin were 11-16 nm and 500,000-1,820,000, respectively. The overall linear conformation of the bovine serum albumin aggregates slightly differed from that of the ovalbumin aggregates since the mass fractal dimensions were found from calculation to be 1.63-1.7 for the bovine serum albumin and 1.36-1.51 for the ovalbumin. The surface property of the aggregates of both proteins was suggested to be similar to that of their native monomer since all the surface fractal dimensions were almost equivalent. The dimensionless Kratky plots of the eluted aggregates indicated that the non-globular conformation of the bovine serum albumin aggregates differs from that of the ovalbumin aggregates. These analyses using size-exclusion chromatography combined with the solution X-ray scattering measurement will be helpful for characterization of the components of the denatured protein aggregation in solution.
在中性 pH 值和低离子强度（10mM 磷酸钠缓冲液，pH 值 6.9）下，采用凝胶渗透色谱法结合小角 X 射线散射测量，对牛血清白蛋白和卵清蛋白在热诱导（80±1℃，1h）下形成的聚集物进行了表征。计算得出牛血清白蛋白洗脱聚集物的回转半径和分子量分别为 9-11nm 和 540,000-820,000。卵清蛋白的分别为 11-16nm 和 500,000-1,820,000。由于从计算得出牛血清白蛋白的质量分形维数为 1.63-1.7，卵清蛋白的质量分形维数为 1.36-1.51，因此牛血清白蛋白聚集物的整体线性构象与卵清蛋白聚集物略有不同。由于所有表面分形维数几乎相等，因此两种蛋白质聚集物的表面性质被认为与它们天然单体的表面性质相似。洗脱聚集物的无量纲 Kratky 图谱表明，牛血清白蛋白聚集物的非球形构象与卵清蛋白聚集物的不同。这些使用凝胶渗透色谱法结合溶液 X 射线散射测量的分析方法将有助于对溶液中变性蛋白质聚集物成分进行表征。
