FOLDING STUDIES OF SINGLE TRP MUTANTS OF RNASE A

我们已经制造了核糖核酸酶A的突变体，其含有单个 色氨酸残基（Trp 92）。 这种突变体具有已知的 性质，可以探测构象变化相关的 热解折叠 我们已经诱导了一个190摄氏度的温度跳跃， 4.5用10 mJ ns脉冲（1.5？m）来自ns OPO; 来自突变体中Trp残基的荧光（在340 nm处）是 在T跳变之前-500 ns至T跳变之后30 ms的时间内进行监测。 T跳 调节蛋白质溶液的静态温度 从250 ℃到550 ℃（通过熔化曲线）。 数据支持 Trp残基周围没有明显的扰动， 时间尺度短于30 ms。此外， 在温度和温度之间荧光强度的增加 变性和GuHCl变性也是明显的，表明 不同的变性机制。

We have made a mutant of Ribonuclease A which contains a single tryptophan residue (Trp92). This mutant has known fluorescent properties which can probe conformational changes associated with thermal unfolding. We have induced a 190C temperature jump in a pH 4.5 solution of Y92W using a 10 mJ ns pulse (1.5 ?m) from the ns OPO; the fluorescence from the Trp residue (at 340 nm) in the mutant is monitored at times from -500 ns before the T-jump to 30 ms after the T-jump. The static temperature of the protein solution was adjusted from 250C to 550C (through the melting curve). The data support that no significant perturbation surrounding the Trp residue is evident on timescales shorter than 30 ms. In addition, a significant difference in the increase of fluorescent intensity between temperature denaturation and GuHCl denaturation is also evident, suggesting differing mechanisms for denaturation.