Bewley T A
Biochemistry. 1977 Jan 25;16(2):209-15. doi: 10.1021/bi00621a008.
The reduction and alkylation of the two disulfide bonds in a preparation of human pituitary growth hormone which had been previously modified by limited proteolysis with the enzyme plasmin have been studied. Quantitative and selective reduction of the carboxyl-terminal disulfide, as well as total reduction of both disulfides, has been achieved in the absence of denaturants. Circular dichroism spectra of the various reduced and reduced-alkylated derivatives have provided sufficient information to allow an estimation of the individual contributions of each disulfide bond to the total optical activity of the protein. These contributions were found to represent a significant portion of the total optical activity between 290 and 250 nm. The carboxyl-termimal bond exhibits negative dichroism with an apparent center near 258 nm ([theta]M,258nm = 2100 deg cm2 dmol-1). By comparison, the contribution of the remaining disulfide is red-shifted to 273 nm, is also negative in sign, and somewhat more intense ([theta]M,273nm = 3200 deg cm2 dmol-1). Circular dichroism measurements have also been used to approximate the rate of reduction of the protein.
对一种经纤溶酶有限度蛋白酶解预先修饰过的人垂体生长激素制剂中的两个二硫键的还原和烷基化反应进行了研究。在不存在变性剂的情况下，实现了羧基末端二硫键的定量和选择性还原以及两个二硫键的完全还原。各种还原和还原烷基化衍生物的圆二色光谱提供了足够的信息，以便估算每个二硫键对蛋白质总光学活性的单独贡献。发现在290至250纳米之间，这些贡献占总光学活性的很大一部分。羧基末端键呈现负二色性，表观中心接近258纳米（[θ]M，258纳米 = 2100度·厘米²·dmol⁻¹）。相比之下，其余二硫键的贡献红移至273纳米，也是负号，且强度稍高（[θ]M，273纳米 = 3200度·厘米²·dmol⁻¹）。圆二色性测量也已用于估算蛋白质的还原速率。
