Fluorescent Proteins Developed for Studies of Inter-subunit and Protein-protein Interactions in the Styrene Catabolic Pathway
Author: Tymesha Bovell
Faculty Supervisor: George Gassner
Department: Chemistry & Biochemistry
Our project explores styrene monooxygenase (SMO), a pivotal enzyme in the Pseudomonas bacteria styrene catabolic pathway, employing fluorescence tagging to elucidate its mechanism. Traditionally identified as a dimer, SMO likely fluctuates between monomer and dimer states during catalysis, engaging with styrene monooxygenase reductase and styrene oxide isomerase (SOI) via transient interactions. We engineered molecular models that append monomeric green (mGreenLantern) and red (mScarlet) fluorescent proteins to SMO, facilitating the observation of these transient states without hindering the enzyme's active site. Following a rigorous structural evaluation, we proceeded with the synthesis of these fusion proteins. Our findings, initially presented at the COSE student research symposium (2023), focused on the integration of linker peptides and ongoing efforts to finalize fluorescent protein attachment. Future studies will leverage stopped flow fluorescence and resonance energy transfer (RET) techniques to monitor SMO's dynamic behavior and interactions, aiming to deepen our understanding of its role within the bacterial catabolic landscape. This research not only sheds light on SMO's operational intricacies but could set the stage for broader applications in biotechnological and pharmaceutical fields.