Figure 5: Fractal assemblies capture and release greater amounts of cargo compared to globular assemblies

Figure 5: Fractal
assemblies capture and release greater amounts of cargo compared to globular
assemblies. Scheme for
the envisioned reversible capture of cargo proteins for the (A) fractal and (B) globular structures. The red stars denote an example cargo
protein (antibody). (C) % protein
capture was measured for 3:2 fractal (assemblies obtained with 3 AtzA-pY: 2
AtzC-SH2), 3:2 GS linker (globular assemblies obtained at the same
stoichiometry with fusion proteins containing long GS-rich linkers), and 3:1
fractal (assemblies obtained with 3 AtzA-pY: 1 AtzC-SH2). The 3:2 fractal
captured more IgG antibody (red bars) and GFP-SH2 (green barss), and degraded
more substrate TCP (purple bar; reflecting the higher capture efficiency of
enzyme DhaA-Sh2) than the 3:2 GS linker assemblies. In addition, 3:2 fractal
released more captured antibody compared to the 3:2 GS linker when incubated
with YopH phosphatase (blue bars). (D,
E) Confocal fluorescence microscopy images
of the 3-component assembly with GFP-SH2 showing the topology of incorporation
of GFP-SH2 in fractal and (F-G) the
incorporation of GFP-SH2 into the globular assemblies. (H-I) the IgG antibody Alexa Fluor 568 incorporation into the
fractal assembly and (J-K) the
incorporation into the globular assembly.

图5：分形组装体相较于球状组装体，可捕获并释放更多的货物分子。本方案展示了针对（A）分形结构与（B）球状结构所设想的货物蛋白可逆捕获流程。红色星号代表示例货物蛋白（抗体）。（C）针对三类组装体测定了蛋白质捕获率：3:2分形组装体（以3份AtzA-pY与2份AtzC-SH2制备得到的组装体）、3:2 GS接头组装体（采用相同化学计量比、融合了富含GS的长接头的蛋白所形成的球状组装体）以及3:1分形组装体（以3份AtzA-pY与1份AtzC-SH2制备得到的组装体）。相较于3:2 GS接头组装体，3:2分形组装体可捕获更多的IgG抗体（红色柱形）与GFP-SH2（绿色柱形），且能降解更多的底物TCP（紫色柱形，反映出酶DhaA-SH2更高的捕获效率）。此外，当与YopH磷酸酶温育时，3:2分形组装体释放的捕获抗体量也多于3:2 GS接头组装体（蓝色柱形）。（D、E）携带GFP-SH2的三组分组装体的共聚焦荧光显微镜图像，展示了GFP-SH2在分形组装体中的掺入拓扑结构；（F~G）为GFP-SH2在球状组装体中的掺入情况。（H~I）为Alexa Fluor 568标记的IgG抗体掺入分形组装体的图像，（J~K）为该抗体掺入球状组装体的图像。