A General, Symmetry-Based Approach for the Assembly of Proteins into Nanoscale Polyhedra.

The assembly of individual protein subunits into large-scale symmetrical structures is widespread in Nature and confers unique biological properties which have potential applications in nano-technology and medicine. While efforts to functionalize and repurpose existing protein complexes have been mainly successful, designing well-defined de novo protein complexes remains an unsolved problem. A major challenge in engineering de novo symmetrical assemblies has been to design interactions between the protein subunits so that they specifically assemble into the desired structure. Prior de novo protein cages have been developed with moderate success, but suffer from a lack of generalizability and require significant computational effort and screening of mutant fusion proteins. The design and optimization of a simple, generalizable approach to designing novel fusion proteins which assemble into cage-like structures will be the subject of this dissertation. We show that by genetically fusing a C4-symmetric coiled-coil to the C-terminus of a C3-symmetric trimeric protein via a short, flexible linker, we can assemble a well-defined 24-subunit protein cage with octahedral symmetry. The flexible nature of these assemblies alleviates the need for rigorous interface modeling, requiring only minimal computation to determine the length of the linker sequence. This is the first de novo designed symmetrical protein complex to incorporate a C4 symmetry element, and we anticipate this method can be applied to a wider variety of proteins and symmetries, which may open up a new avenue of research into designer protein cages with unique, built-in functionalities.PHDChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/120840/1/sciore_1.pd

Symmetryâ Directed Selfâ Assembly of a Tetrahedral Protein Cage Mediated by de Novoâ Designed Coiled Coils

The organization of proteins into new hierarchical forms is an important challenge in synthetic biology. However, engineering new interactions between protein subunits is technically challenging and typically requires extensive redesign of proteinâ protein interfaces. We have developed a conceptually simple approach, based on symmetry principles, that uses short coiledâ coil domains to assemble proteins into higherâ order structures. Here, we demonstrate the assembly of a trimeric enzyme into a wellâ defined tetrahedral cage. This was achieved by genetically fusing a trimeric coiledâ coil domain to its C terminus through a flexible polyglycine linker sequence. The linker length and coiledâ coil strength were the only parameters that needed to be optimized to obtain a high yield of correctly assembled protein cages.Geometry lesson: A modular approach for assembling proteins into largeâ scale geometric structures was developed in which coiledâ coil domains acted as â twist tiesâ to facilitate assembly. The geometry of the cage was specified primarily by the rotational symmetries of the coiled coil and building block protein and was largely independent of protein structural details.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/138862/1/cbic201700406_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/138862/2/cbic201700406.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/138862/3/cbic201700406-sup-0001-misc_information.pd

Heavy Metals in Dianshan Lake

In this study, the sediment of the Dianshan Lake, one of the major freshwater sources for Shanghai, was analyzed at varying depths to determine its historical environmental toxicity profile. Six heavy metal elements (Cd, Cr, Pb, Zn, Hg, and As) were analyzed, using a three step Sequential Extraction Procedure to determine the speciation of each metal, which gives a more accurate profile on their mobility and bioavailability(toxicity). The order of mobility was found to be Cd > Pb > Zn > Cr > Hg > As. Levels of mobile species of Cr, Zn, Hg, and As were found to increase in the past decade, while Cd and Pb have remained constant. Peaks in mobile or total species concentration for Cr, Pb, and As are observable at 9-13 cm depth, corresponding to China's rapid industrialization in the 1980's