Hahn T R, Song P S
Biochemistry. 1982 Mar 16;21(6):1394-9. doi: 10.1021/bi00535a045.
The hydrogen-tritium-exchange measurements on phytochrome have been performed to detect the conformational differences between the red-absorbing (Pr) and the far-red absorbing (Pfr) forms of phytochrome. The large and small Pfr molecules revealed more exchangeable protons that did the corresponding Pr molecules by 96 and 70 protons, respectively. These results suggest that the Pr leads to Pfr phototransformation is accompanied by an additional exposure of the peptide chains in the Pfr molecule. Of 1682 theoretically exchangeable hydrogens in undegraded phytochrome, only 442 (26%) and 346 (21%) protons were found to be exchangeable (excluding instantaneously exchangeable protons that cannot be determined by the present method). Thus, the phytochrome protein appears to be compact and highly folded. The kinetic analyses of the tritium exchange-out curves indicate that two kinetically different groups are responsible for the conformational differences between the Pr and Pfr forms of phytochrome. These components are due to (1) the exposure of hydrogen-bonded peptide segments (alpha helix and/or beta-pleated sheet) in the chromophore vicinity of Pfr and (2) the exposure of hydrogen-bonded peptide segments on the chromophore peptide domain as well as on the chromophore-free tryptic domain of undegraded phytochrome.
已对光敏色素进行了氢 - 氚交换测量，以检测光敏色素的红光吸收型（Pr）和远红光吸收型（Pfr）之间的构象差异。大的和小的Pfr分子分别比相应的Pr分子显示出更多可交换的质子，多出96个和70个质子。这些结果表明，Pr向Pfr的光转化伴随着Pfr分子中肽链的额外暴露。在未降解的光敏色素中理论上可交换的1682个氢中，仅发现442个（26%）和346个（21%）质子是可交换的（不包括用本方法无法测定的瞬时可交换质子）。因此，光敏色素蛋白似乎是紧密且高度折叠的。氚交换出曲线的动力学分析表明，两个动力学上不同的基团导致了光敏色素Pr和Pfr形式之间的构象差异。这些组分归因于：（1）Pfr发色团附近氢键结合的肽段（α螺旋和/或β折叠片）的暴露；（2）未降解的光敏色素发色团肽结构域以及无发色团的胰蛋白酶消化结构域上氢键结合的肽段的暴露。
