This project is funded jointly by the Biomolecular Systems and Cellular Systems clusters.
A complex array of photoreceptors coordinates the response of both prokaryotes and eukaryotes to their ambient light environment. One of the most influential is the phytochrome superfamily, a large and diverse group of photoreversible photoreceptors that use a bilin (or linear tetrapyrrole) chromophore for light detection. Phytochromes sense red (R) and far-red light (FR) through two relatively stable conformations, a R-absorbing Pr form and a FR-absorbing Pfr form. By photointerconverting between Pr and Pfr, phytochromes act as light-regulated switches in various signaling cascades. Despite their agricultural importance and evolutionary conservation among plants and microorganisms, it is not fully understood how phytochromes photoconvert between Pr and Pfr at the molecular level or how this switch initiates the responses under phytochrome control. The prior NSF-funded studies provided a major breakthrough toward determining how phytochromes work at the atomic level by generating the first 3-D structure of the bilin-binding domain of a phytochrtome as Pr. This structure conclusively determined the chemistry and conformation of the bilin, revealed that the chromophore pocket is uniquely folded into a rare figure-of-eight knot, identified a heretofore unknown dimerization contact, and provided important clues for how plant Phys arose from their microbial progenitors. This project continues the structural analysis to determine how phytochromes photoconvert from Pr to Pfr, how phytochromes evolved, and how organisms assemble knotted proteins in general. In particular, the project will determine the 3-D structures of larger fragments of phytochromes, toward the eventual goal of deducing an entire phytochrome structure, (2) determine the first 3-D structure of the enigmatic Pfr form by exploiting novel naturally-occurring phytochromes that are more amenable to structural studies, and (3) identify key amino acids that participate in Pr to Pfr photochemistry, dimerization, knot assembly, and ultimately signal transmission by various biochemical and biophysical assays or in vivo systems that directly respond to phytochrome signals. This project will help elucidate how microorganisms and plants sense their light environment, which could have important ramifications for understanding microbial ecosystems, the control of important microbial pathogens, and for the development of new strategies to improve the productivity of both food and biofuel crops.
Broader Impacts: The project involves research training of postdoctoral and graduate students. It will also enhance scientific infrastructure via a cooperative college/university arrangement for the training of minority undergraduate students in modern molecular techniques. This student training program will provide summer research experience to undergraduate students from University of West Alabama.
