Bioengineering and Analysis of Near-Infrared Absorbing Optogenetic Rhodopsins

Speaker: Gaoxiang Mei

When: October 29, 2019 (Tue), 02:00PM to 03:00PM (add to my calendar)
Location: SCI 352

This event is part of the Preliminary Oral Exam.

Examining Committee: Kenneth Rothschild, Shyam Erramilli, Claudio Chamon, Robert Carey

Abstract:

Optogenetics is a biological technique that allows researchers to control neuronal activity with light. It has been applied in a variety of important fields such as probing the role of individual neurons in the brain. Neuronal control in optogenetics is achieved using optogenetic actuators like microbial rhodopsins. However, effective in vivo optogenetics is in many cases severely limited due to the strong absorption and scattering of visible light by biological tissues. Recently, a combination of opsin site-directed mutagenesis and analog retinal substitution has produced variants of microbial rhodopsins which absorb maximally in the near-infrared (NIR). One class of such microbial rhodopsins discovered in marine proteobacteria, called proteorhodopsins (PRs), have diverse functions including serving as light-driven proton pumps. Although PRs have been engineered for use in a variety of biotechnological applications, the molecular mechanism underlying their function is not fully understood. Better understanding of their mechanism would facilitate bioengineering of PRs’ properties such as the wavelength of absorption for specific applications. This talk will first cover resonance Raman and FTIR-difference spectroscopy that helped elucidate the molecular mechanism of bacteriorhodopsin, the first microbial rhodopsin to be studied. Then latest results from UV-Visible-NIR absorption and resonance Raman spectroscopy will be presented on green absorbing proteorhodopsin (GPR) and its mutants, which exhibit some similarities with bacteriorhodopsin. Utilizing the red-shifted double mutant D212N/F234S and the substitution of native A1 retinal with analog retinal 3-methylamino-16-nor-1,2,3,4-didehydroretinal (MMAR), a dramatic red-shift of approximately 200 nm is achieved with this GPR-DNFS:MR. The talk will end with an outline of future work on other optogenetic proteins such as the florescent voltage sensing archaerhodopsin-3 (AR3) and its mutants.