PROTEIN BACKBONE STRUCTURE CHANGES THRU ACTIVE VALINE RESIDUES IN PHOTOREACTION

来自X射线、中子衍射和电子显微镜的证据 表明Fhelix在BR光循环期间倾斜，打开 Sb区对质子的进入和BR的G-螺旋 在质子运输过程中经历了显著的结构变化。 巧合的是，BR中的21个Valine残基中有10个存在于这两个残基中 螺旋，F中7个(Val167、173、177、179、180、187和188)，G中3个 (VA1210、213和217)。根据亨德森的BR结构模型，681 有两个直接键合的Valine对(Vall79和VaI180，VaI187 和VaI188)在富含缬氨酸的F-螺旋上。在每一种情况下， 羰基在空间上是紧密的(-3Ao)。它们之间的距离 F、A、Va1210和V11213和Va1213中的VaI177和VaI179对 G中的Va1217约为4.5A~5。此外，~(13)CCP/MAS核磁共振 光适应(LA)态与M。 [1-13CVal]，[15N-Prol-BR]的光中间体表明该化学物质 21个缬氨酸残基中至少有两个的移位前移了at 至少3.5ppm，这表明 这些Valine残留物。这意味着这些Valine残基是 在BR光循环期间处于活动状态。Valine 49的选择性观察， 69和199通过异核-偶极重新耦合(REDOR)已经消除 可能是三个不存在于 F或G螺旋是导致观察到的变化的原因 核磁共振差谱。因此，这是值得的 调查F和F中的Valine残基是否发生这些变化 在LA-+M转换中的G螺旋。同核偶极子再耦合 (情节剧)实验在较短的混合时间内按顺序进行 仅将相距约3A的两个Valine对重新连接 更长的混合时间以关联F和G中的三个valine对 螺旋，它们被进一步分开。结果表明，G。 Helix可能经历了早期M和ML状态的变化，原因是 它与视网膜发色团的直接结合及SB的作用 反离子残留物Asp212。这些变化在 M[.-~M[，，过渡。然而，F螺旋相对惰性。 BR光循环前半部分的光反应。

Evidence from x-ray, neutron diffraction and electron microscopy suggests that the Fhelix is tilted during the bR photocycle, opening the entrance of the SB region to protons and the G-helix of bR undergoes significant structural changes during the proton transport. Coincidently, 10 of the 21 valine residues in bR populate these two helices, 7 in F (Val 167, 173, 177, 179, 180, 187 and 188) and 3 in G (Va1210, 213, and 217). From Henderson's structural model for bR,681 there are two directly bonded valine-pairs (Vall79 and VaI180, VaI187 and VaI188) in the valine rich F-helix. In each of them, the carbonyls are close in space (-3 AO). The distances between the valine pairs VaI177 and VaI179 in F, A Va1210 and V11213, andVa1213 and Va1217 in G are about4.5 A to 5 Furthermore, the 13CCP/MAS NMR difference spectrum between the light-adapted (LA) state and M. photointermediate of [1-13CVal], [15N-Prol-bR shows that the chemical shifts of at least two of the 21 valine residues move upfield by at least 3.5 ppm, an indication of significant structural changes near these valine residues. This implies that these valine residues are active during the bR photocycle. Selective observation of valines 49, 69 and 199 by heteronucleaf-dipolar recoupling (REDOR) has eliminated the possibility that the three valine residues, which are not in either the F or G helices, are responsible for the changes observed in the NMR difference spectrum. Therefore, it is worthwhile to investigate if these changes occur on the valine residues in the F and G helices in the LA-+M. transition. Homonuclear dipolar recoupling (MELODRAMA) experiments were performed at a short mixing time in order to recouple only the two valine pairs located about 3 A apart and at longer mixing time to correlate the three valine pairs in the F and G helices, which are further separated. The results indicate that the G helix likely undergoes changes in the early M and ML states, due to its direct bonding to the retinal chromophore and the role of the SB counterion residue Asp212. These changes disappear during the M[.-~M[,, transition. However, the F helix is relatively inert to the photoreactions in the first half of the bR photocycle.