A Novel Synthetic Yeast for Enzymatic Biodigester Pretreatment

Lignin, a complex organic polymer, is a major roadblock to the efficiency of biofuel conversion as it both physically blocks carbohydrate substrates and poisons biomass degrading enzymes, even if broken down to monomer units. A pretreatment process is often applied to separate the lignin from biomass prior to biofuel conversion. However, contemporary methods of pretreatment require large amounts of energy, which may be economically uncompelling or unfeasible. Taking inspiration from several genes that have been isolated from termites and fungi which translate to enzymes that degrade lignin, we want to establish a novel “enzymatic pretreatment” system where microbes secrete these enzymes to degrade lignocellulosic biomass. We incorporated the following genes into yeast vectors: laccase, lignin peroxidase, and alpha-keto-reductase from Reticulitermes flavipes; versatile peroxidase from Colletotrichum fioriniae PJ7; manganese peroxide from Heterobasidion irregulare TC 32-1; and tyrosinase from Agaricus bisporus. These vectors code for fusion proteins with yeast secretion tags at the end of each enzyme gene, fluorescent protein tags at the beginning, as well as standardized restriction sites for synthetic biology manipulation. Furthermore, we designed an additional vector to contain our genetically modified yeast using an oxygen-repressed killswitch. We expect that transformants with our construct will be able to secrete said enzymes and contribute to lignin degradation if added to a biomass slurry. Future studies may focus on constructing a prototype bioreactor system and optimizing which combination of enzymes lead to the most efficient biofuel production

Harnessing Notch Signaling for Biomaterial Scaffold-based Bone Regeneration

Bone fracture has recently become prevalent, especially with an increasingly aging population. Current bone grafts procedures, including autografts and allografts, are hindered by multiple factors, such as limited supplies and inconsistent bone healing. Scaffold-based bone tissue engineering emerges as a prospective strategy to aid in bone regeneration through delivery of growth factors such as bone morphogenic proteins (BMPs). However, the use of BMPs suffers from several drawbacks such as protein instability and immunogenicity. Therefore, there exists a great need for the development of novel therapies to promote bone healing. Notch signaling, a pathway critical for cell-fate determination has been shown to regulate osteogenesis, which suggests the potential of targeting Notch to enhance bone repair. The long-term goal of this work is to develop biomaterial-based regenerative technologies to induce bone regeneration by fine-tuning Notch signaling. In this study, a three-dimensional (3D) porous scaffold system was fabricated from biodegradable poly(lactide-co-glycolide) (PLGA) to mimic structural and mechanical properties of native bone using a microsphere sintering technology. In vitro studies were conducted to evaluate the effects of notch inhibition via a γ-secretase inhibitor DAPT on osteoblast responses. When the DAPT was added during the 2D culture on tissue culture polystyrene (TCPS), the osteoblast mineral deposition was significantly enhanced. Intriguingly, the enhancement was more pronounced on the 3D PLGA scaffolds, which may be attributed to the increased cell-cell contact in the 3D culture environment. Current efforts are focused on scaffold-based modulation of Notch signaling with both quantitative and temporal precision for enhanced osteogenesis