Abstract
Cellulose is one of the most abundant biopolymers for food, fiber and fuel. This versatile
fiber is made by plant cellulose synthase, a plant membrane protein with a structural architecture
that is not fully understood. PpCesA5 from an early moss plant Physcomitrium patens has been the
protein of interest due to PpCesA5-specific homo-oligomerization. This oligomerization can form
trimers and a hexamer of CesA trimers called cellulose synthase complexes (CSCs) found in moss
plants. Our 3 Ã… PpCesA5 trimer cryo-EM structure showed structural similarities to higher plant
CesAs and bacterial CesAs. This work is the first cryo-EM structure of the Center for Nanophase
Materials Sciences at ORNL. Concerted flexible movement was discovered between PpCesA5
trimerization axis formed by the plant conserved region (PCR) and a putative phosphate ion
coordinated by PpCesA5 PCR. Lastly, a novel tether pin region was revealed that tethers the
intrinsically disordered N-terminal domain (NTD) of PpCesA5 to the protein main body. The
structural conformation of the tethered NTD would infer loop formation, but cryo-EM was unable
to detect intrinsically disordered regions in PpCesA5. Small-angle neutron scattering (SANS)
showed that the NTD forms extended loops formed by the tether pin. This flexible protein region
gives new leads onto CSC assembly involving the tether pin. This PpCesA5 NTD region was
isolated and further analyzed using small-angle X-ray scattering (SAXS). Contrary to previous
knowledge on NTD dimerization, this presentation will show that NTD formed extended
monomers, further supporting an extended NTD conformation in PpCesA5. Additional work is
underway to build a nanodisc-bound DNA origami corral that can cluster PpCesA5 trimers into
CSCs in vitro. GraFix chemical fixation of AtCesA1 CatD trimer was used as a workhorse to create
new avenues in in vitro CesA stability. Overall, this research reveals new oligomerization
mechanisms and informs rational design of biofuels and new biomaterials.
