Phanerozoic levels of atmospheric oxygen relate to the burial histories of organic carbon and pyrite sulfur. The sulfur cycle remains poorly constrained, however, leading to concomitant uncertainties in O 2 budgets. Here we present experiments linking the magnitude of fractionations of the multiple sulfur isotopes to the rate of microbial sulfate reduction. The data demonstrate that such fractionations are controlled by the availability of electron donor (organic matter), rather than by the concentration of electron acceptor (sulfate), an environmental constraint that varies among sedimentary burial environments. By coupling these results with a sediment biogeochemical model of pyrite burial, we find a strong relationship between observed sulfur isotope fractionations over the last 200 Ma and the areal extent of shallow seafloor environments. We interpret this as a global dependency of the rate of microbial sulfate reduction on the availability of organic-rich sea-floor settings. However, fractionation during the early/mid-Paleozoic fails to correlate with shelf area. We suggest that this decoupling reflects a shallower paleoredox boundary, primarily confined to the water column in the early Phanerozoic. The transition between these two states begins during the Carboniferous and concludes approximately around the Triassic-Jurassic boundary, indicating a prolonged response to a Carboniferous rise in O 2 . Together, these results lay the foundation for decoupling changes in sulfate reduction rates from the global average record of pyrite burial, highlighting how the local nature of sedimentary processes affects global records. This distinction greatly refines our understanding of the S cycle and its relationship to the history of atmospheric oxygen.Phanerozoic oxygen | sulfate-reducing bacteria T he marine sedimentary sulfur isotope record encodes information on the chemical and biological composition of Earth's ancient oceans and atmosphere (1, 2). However, our interpretation of the isotopic composition of S-bearing minerals is only as robust as our understanding of the mechanisms that impart a fractionation. Fortunately, decades of research identify microbial sulfate reduction (MSR) as the key catalyst of the marine S cycle, both setting the S cycle in motion and dominating the massdependent fractionation preserved within the geological record (1, 3, 4). Despite the large range of S-isotope variability observed in biological studies (4-6), attempts to calibrate the fractionations associated with MSR are less mechanistically definitive (7, 8) than analogous processes influencing the carbon cycle (9, 10). What is required is a means to predict S isotope signatures as a function of the physiological response to environmental conditions (e.g., reduction-oxidation potential).Microbial sulfate reduction couples the oxidation of organic matter or molecular hydrogen to the production of sulfide, setting in motion a cascade of reactions that come to define the biogeochemical S cycle. In modern marine sediments, sulfide i...

Microbial sulfate reduction has governed Earth's biogeochemical sulfur cycle for at least 2.5 billion years. However, the enzymatic mechanisms behind this pathway are incompletely understood, particularly for the reduction of sulfite-a key intermediate in the pathway. This critical reaction is performed by DsrAB, a widespread enzyme also involved in other dissimilatory sulfur metabolisms. Using in vitro assays with an archaeal DsrAB, supported with genetic experiments in a bacterial system, we show that the product of sulfite reduction by DsrAB is a protein-based trisulfide, in which a sulfite-derived sulfur is bridging two conserved cysteines of DsrC. Physiological studies also reveal that sulfate reduction rates are determined by cellular levels of DsrC. Dissimilatory sulfate reduction couples the four-electron reduction of the DsrC trisulfide to energy conservation.

These data reveal complexity in the sulfate concentration-fractionation relationship. 42Sulfur isotope fractionation during sulfate reduction relates to environmental sulfate 43 concentrations but also to strain-specific physiological parameters such as the affinity of sulfate-44 reducing microorganisms for sulfate and electron donors. Previous studies have suggested that 45 the relationship between sulfate concentration and isotope fractionation is best fit with a MM fit. 46suggested We present a simple model, grounded in the physiology of sulfate reduction, in which 47 the ratio of MM relationships for sulfate and electron donor uptake produces the relationships 48 seen in experimental studies: a MM relationship with sulfate concentration, and a hyperbolic 49 relationship with growth rate. 50Since both environmental and biological factors influence the fractionation recorded in 51 geological samples, understanding their relationship is critical to interpreting the sulfur isotope 52 record. As the acquisition machinery for sulfate and electron acquisition has been subject to 53 selective pressure over Earth history, its evolution may complicate efforts to uniquely reconstruct 54 ambient sulfate concentrations from a single sulfur isotopic composition. 55Patterns of SRB S-isotope fractionation 2 56