GSA Annual Meeting in Phoenix, Arizona, USA - 2019

Paper No. 104-4
Presentation Time: 9:00 AM-6:30 PM


GARCIA, Amanda, Department of Molecular and Cellular Biology, University of Arizona, 1007 E Lowell St, Tucson, AZ 85721, MCSHEA, Hanon, Department of Earth System Science, Stanford University, Stanford, CA 94305, KOLACZKOWSKI, Bryan, Department of Microbiology and Cell Science, University of Florida, Gainesville, FL 32611 and KACAR, Betul, Department of Astronomy and Steward Observatory, University of Arizona, Tucson, AZ 85721; Department of Molecular and Cellular Biology, University of Arizona, 1007 E Lowell St, Tucson, AZ 85721

Nitrogen, an essential element for life, is made bioavailable by nitrogenase metalloenzymes that catalyze the reduction of N2 to NH3. Nitrogenases have an ancient evolutionary history, likely having supplied the bulk of fixed nitrogen to the biosphere since the Archean. The most common form of modern nitrogenase binds a Mo-containing active-site metal cofactor and rarer forms bind V- or Fe-only cofactors. Putative marine Mo scarcity prior to atmospheric oxygenation at ~2.3 Ga has led to the suggestion that Fe-nitrogenases may have been ancestral due to metal-availability constraints on early nitrogenase evolution. More recent phylogenetic reconstructions have instead indicated that the evolution of Mo-nitrogenases preceded that of V- and Fe-nitrogenases. However, ~3.2-Ga isotopic signatures of biological nitrogen fixation support an earlier origin of nitrogenase, preceding age estimates of both Mo-nitrogenase and the onset of marine Mo availability. Thus, the precise nature of the coevolution of nitrogenase metal usage and marine geochemistry remains uncertain. We explored the evolution of nitrogenase metal usage by reconstructing ancestral nitrogenase protein sequences and structures in silico, as well as by investigating the presence of nitrogenase cofactor biosynthetic genes in modern taxa. Ancestors were inferred from a maximum likelihood phylogeny of 284 modern sequences along an evolutionary transect between Mo- and V-/Fe-nitrogenases. Ancestral variant sequences were also inferred from alternate phylogenetic topologies, as well as by Bayesian sampling of maximum likelihood site posterior probabilities. We analyzed both sequence and structural features of ancestral nitrogenases previously used to classify modern nitrogenase metal dependence. Though these analyses suggest that oldest ancestral nitrogenases resemble modern Mo-nitrogenases, phylogenetic evidence indicates that early nitrogenases may not have been associated with biosynthetic mechanisms necessary for Mo-dependence. Future investigations of ancient nitrogenase metal usage should be augmented by laboratory restorations of ancestral enzymes.