Chumachenko Iu V, Sytnik A I
Ukr Biokhim Zh (1978). 1986 Sep-Oct;58(5):32-7.
It is shown by the fluorimetric analysis that with the 1,2 M MgCl2-induced dissociation of rabbit muscle aldolase the tertiary structure of the resulted protomers (subunits) remains practically unchanged. Significant changes in the protomeric enzyme are provoked by subsequent addition of urea up to the concentration of 2,3 M, and are, evidently, manifested in a significant decrease in regularity of the hydrophobic part of aldolase and in possible transition of its Trp-147 into more polar environment. This transition is reflected in the longwave shift of the protein fluorescence maximum (lambda max) by 13 nm (from 320 to 333 nm). But the joint action of MgCl2 and urea does not lead to complete unfolding of the resulted protomeric enzyme. More deep structural alterations in the subunits occur on acidic dissociation, and lambda max shift in this case reaches 342 nm. Structural changes caused by MgCl2 and urea are concomitant with the increase of fluorescence quenchibility with NADH. Here a short-wave lambda max shift, being usually observed in native aldolase fluorescence quenching, is not registered. This mean that the photoselection of protein fluorophores does not occur. The results thus obtained produce an evidence that oligomerization endows aldolase protomers with enhanced stability.
荧光分析表明，在1.2M MgCl₂诱导兔肌肉醛缩酶解离时，所得原体（亚基）的三级结构实际上保持不变。随后添加高达2.3M浓度的尿素会引发原体酶的显著变化，显然，这表现为醛缩酶疏水部分的规则性显著降低，以及其色氨酸-147可能向极性更强的环境转变。这种转变反映在蛋白质荧光最大值（λmax）的长波位移上，位移了13nm（从320nm到333nm）。但是MgCl₂和尿素的联合作用并不会导致所得原体酶完全展开。在酸性解离时，亚基会发生更深入的结构改变，此时λmax位移达到342nm。MgCl₂和尿素引起的结构变化伴随着与NADH荧光猝灭能力的增加。在此，通常在天然醛缩酶荧光猝灭中观察到的短波λmax位移并未出现。这意味着蛋白质荧光团的光选择并未发生。由此获得的结果证明寡聚化赋予醛缩酶原体更高的稳定性。
