Public Abstract

DE-FG02-08ER15968: STRUCTURE, FUNCTION, AND REGULATION OF THE NDH-1 COMPLEXES IN CYANOBACTERIA

Award Status: Active

Institution: Oklahoma State University, Stillwater, OK
UEI: NNYDFK5FTSX9
PM: Brown, Katherine

Most Recent Award Date: 06/09/2026
Number of Support Periods: 18
PI: Burnap, Robert

Current Budget Period: 07/15/2026 - 07/14/2027
Current Project Period: 07/15/2024 - 07/14/2027

Public Abstract

The overall objective is better to understand the uptake of CO2 by cyanobacteria for photosynthesis. A continuation of a structure/function project is proposed to understand NDH-1 complexes that concentrate CO2 inside cyanobacterial cells. The cyanobacterial NDH-1 complexes exhibit a modular structural organization consisting of a redox energy-transducing core module fitted with additional paralogous submodules conferring different physiological functions depending upon the specific proteins associated with it. The NDH-1 complexes associated with CO2 uptake (Cup) proteins use redox energy to hydrate CO2, providing bicarbonate to the carboxysome and, in the process, contributing to cyclic electron flow (CEF). Despite its critical role in the CCM and even with a recent cryo-EM structure of the NDH-1/Cup complex, the molecular details of how NDH-1/Cup energizes conversion of CO2 to bicarbonate. There are two broad objectives: • Elucidate the Zn2+-containing active site and how it facilitates the direct hydration of CO2 into bicarbonate. • Understand how the CO2-hydration reaction is coupled to the energy transduction processes in the bioenergetic machinery of the NDH-1 complex. The project will utilize and improve structural information as a foundation for computational studies and to guide the mutagenesis of CO2 uptake modules. It builds on a newly engineered cyanobacterial mutant that over-expresses one form of the CO2 hydration complex, NDH-1(4), without any other form of the complex, including the respiratory NDH-1(1/2) complex. This mutant will allow an unprecedented ability to probe its mechanistic features without complicating parallel activities of the complex's other paralogous and normally co-expressed forms. Functional analysis will be combined with structural and computational methods to refine and test hypotheses to elucidate the highly unusual configuration of the Zn2+ active side and evaluate the prediction of proton-conduction pathways connecting the active site of CO2 hydration to the proton-pumping apparatus of the transmembrane antiporter-like subunits. The rationale for this work is that the acquisition of inorganic carbon often limits photosynthesis in natural systems, and this limitation is likely to be even more acute in engineered systems that may have proportionally higher demands for inorganic carbon. The mechanism of energetic coupling to CO2-hydration chemistry remains completely unresolved. The reaction mechanism could provide ideas for biomimetic devices aimed at CO2 capture. The project will also help expand an outreach initiative for Oklahoma public school educators, extending its impact to a wide demographic of students within the public school system. This effort builds on a pioneering program for high school teachers of the past 12 years. In a novel extension of the program, we will leverage our proficiency in bioinformatics and computational analysis to create exercises that enhance universally relevant computer skills related to bioinformatics. This curriculum development will guide teachers in integrating this advanced knowledge into their lesson plans, aligning with state educational standards.

The overall objective is better to understand the uptake of CO2 by cyanobacteria for photosynthesis. A continuation of a structure/function project is proposed to understand NDH-1 complexes that concentrate CO2 inside cyanobacterial cells. The cyanobacterial NDH-1 complexes exhibit a modular structural organization consisting of a redox energy-transducing core module fitted with additional paralogous submodules conferring different physiological functions depending upon the specific proteins associated with it. The NDH-1 complexes associated with CO2 uptake (Cup) proteins use redox energy to hydrate CO2, providing bicarbonate to the carboxysome and, in the process, contributing to cyclic electron flow (CEF). Despite its critical role in the CCM and even with a recent cryo-EM structure of the NDH-1/Cup complex, the molecular details of how NDH-1/Cup energizes conversion of CO2 to bicarbonate. There are two broad objectives:

• Elucidate the Zn2+-containing active site and how it facilitates the direct hydration of CO2
into bicarbonate.
• Understand how the CO2-hydration reaction is coupled to the energy transduction processes
in the bioenergetic machinery of the NDH-1 complex.

The project will utilize and improve structural information as a foundation for computational studies and to guide the mutagenesis of CO2 uptake modules. It builds on a newly engineered cyanobacterial mutant that over-expresses one form of the CO2 hydration complex, NDH-1(4), without any other form of the complex, including the respiratory NDH-1(1/2) complex. This mutant will allow an unprecedented ability to probe its mechanistic features without complicating parallel activities of the complex's other paralogous and normally co-expressed forms. Functional analysis will be combined with structural and computational methods to refine and test hypotheses to elucidate the highly unusual configuration of the Zn2+ active side and evaluate the prediction of proton-conduction pathways connecting the active site of CO2 hydration to the proton-pumping apparatus of the transmembrane antiporter-like subunits. The rationale for this work is that the acquisition of inorganic carbon often limits photosynthesis in natural systems, and this limitation is likely to be even more acute in engineered systems that may have proportionally higher demands for inorganic carbon. The mechanism of energetic coupling to CO2-hydration chemistry remains completely unresolved. The reaction mechanism could provide ideas for biomimetic devices aimed at CO2 capture. The project will also help expand an outreach initiative for Oklahoma public school educators, extending its impact to a wide demographic of students within the public school system. This effort builds on a pioneering program for high school teachers of the past 12 years. In a novel extension of the program, we will leverage our proficiency in bioinformatics and computational analysis to create exercises that enhance universally relevant computer skills related to bioinformatics. This curriculum development will guide teachers in integrating this advanced knowledge into their lesson plans, aligning with state educational standards.