Institute of Biophysics and Physical Biochemistry and Centre of Magnetic Resonance in Chemistry and Biomedicine, University of Regensburg, Regensburg, Germany.
PLoS One. 2024 Apr 18;19(4):e0301579. doi: 10.1371/journal.pone.0301579. eCollection 2024.
We present here the solution structures of the protein thioredoxin-1 from Plasmodium falciparum (PfTrx-1), in its reduced and oxidized forms. They were determined by high-resolution NMR spectroscopy at 293 K on uniformly 13C-, 15N-enriched, matched samples allowing to identification of even small structural differences. PfTrx-1 shows an α/β-fold with a mixed five-stranded β-sheet that is sandwiched between 4 helices in a β1 α1 β2 α2 β3 α3 β4 β5 α4 topology. The redox process of the CGPC motif leads to significant structural changes accompanied by larger chemical shift changes from residue Phe25 to Ile36, Thr70 to Thr74, and Leu88 to Asn91. By high-field high-pressure NMR spectroscopy, rare conformational states can be identified that potentially are functionally important and can be used for targeted drug development. We performed these experiments in the pressure range from 0.1 MPa to 200 MPa. The mean combined, random-coil corrected B1* values of reduced and oxidized thioredoxin are quite similar with -0.145 and -0.114 ppm GPa-1, respectively. The mean combined, random-coil corrected B2* values in the reduced and oxidized form are 0.179 and 0.119 ppm GPa-2, respectively. The mean ratios of the pressure coefficients B2/B1 are -0.484 and -0.831 GPa-1 in the reduced and oxidized form respectively. They differ at some points in the structure after the formation of the disulfide bond between C30 and C33. The thermodynamical description of the pressure dependence of chemical shifts requires the assumption of at least three coexisting conformational states of PfTrx-1. These three conformational states were identified in the reduced as well as in the oxidized form of the protein, therefore, they represent sub-states of the two main oxidation states of PfTrx-1.
我们在此介绍恶性疟原虫（Plasmodium falciparum）硫氧还蛋白-1（PfTrx-1）在还原和氧化两种形式下的溶液结构。这些结构是通过在 293 K 下使用高分辨率 NMR 光谱法，对均进行了 13C、15N 均匀标记和匹配的样品进行测定得到的，这使得即使是很小的结构差异也能够被识别。PfTrx-1 呈现出一个具有混合五股β-折叠的α/β折叠结构，其中β折叠夹在 4 个螺旋之间，拓扑结构为β1α1β2α2β3α3β4β5α4。CGPC 基序的氧化还原过程导致了显著的结构变化，同时残基 Phe25 到 Ile36、Thr70 到 Thr74 和 Leu88 到 Asn91 的化学位移变化也更大。通过高场高压 NMR 光谱学，可以鉴定出潜在功能重要的稀有构象状态，并可用于靶向药物开发。我们在 0.1 MPa 至 200 MPa 的压力范围内进行了这些实验。还原和氧化形式的硫氧还蛋白的平均组合、随机卷曲校正后的 B1值非常相似，分别为-0.145 和-0.114 ppm GPa-1。还原和氧化形式的平均组合、随机卷曲校正后的 B2值分别为 0.179 和 0.119 ppm GPa-2。还原和氧化形式的 B2/B1 压力系数比分别为-0.484 和-0.831 GPa-1。在 C30 和 C33 之间形成二硫键后，结构的某些点上会出现差异。化学位移对压力依赖性的热力学描述需要假设 PfTrx-1 至少存在三种共存的构象状态。在蛋白质的还原和氧化形式中都鉴定出了这三种构象状态，因此，它们代表了 PfTrx-1 的两种主要氧化状态的亚状态。
