The uptake and interconversion of inorganic carbon are major life processes that are necessary to sustain growth and reproduction of organisms, processes that in plants precede photosynthetic CO2 fixation. Using a prokaryote, the marine, unicellular, nitrogen-fixing cyanobacterium Synechococcus sp.SF1 as a model system for a photo-autotrophic cell, will enable us to study inorganic carbon-utilizing reactions in the same cell and compare these reactions with those of other nitrogen-fixing organisms, with higher plants, and with mammalian lung, bone and kidney. The experiments outlined in this proposal, once completed, will provide the following information: a) The uptake and kinetics of CO2 and bicarbonate utilization in Synechococcus. Formation of intracellular pools of inorganic carbon. Determination of their pool size by the CO2 fixation depletion method; b) the role of K+ and Na+ in the uptake of inorganic carbon and the regulation of the pH of the cytosol in dark and light conditions; c) The role of the electrochemical potential of protons in dark/light transients of Synechococcus cells as it effects inorganic carbon uptake and nitrogen fixation; d) The mechanism of O2 exchange in dark and light conditions, using bicarbonate as a carbon source; e) The role of carbonic anhydrase in the turnover of inorganic carbon in the cell during the light period; f) Purification of carbonic anhydrase from Synechococcus and comparison with other carbonic anhydrases from plants and animals. Significance: The study of the CO2/HCO3- exchange and characterization of carbonic anhydrases in Synechococcus will help understand the closely related processes catalyzed by human carbonic anhydrases I,II, and III in red cells, in capillary endothelium of lungs, and in red skeletal muscle. The processes of CO2 gas exchange during respiration and CO2 fixation in the more primitive marin cyanobacteria that are related to the regulation of the pH of cells, will provide a more thorough understanding of the processes of acid/base homeostasis in human organs of stomach and uterus.
