Sulfur Isotope Effects of Dissimilatory Sulfite Reductase.

William D Leavitt,Inês A C Pereira,André A Santos,David T Johnston,Alexander S Bradley

doi:10.3389/fmicb.2015.01392

Abstract

The precise interpretation of environmental sulfur isotope records requires a quantitative understanding of the biochemical controls on sulfur isotope fractionation by the principle isotope-fractionating process within the S cycle, microbial sulfate reduction (MSR). Here we provide the only direct observation of the major (34S/32S) and minor (33S/32S, 36S/32S) sulfur isotope fractionations imparted by a central enzyme in the energy metabolism of sulfate reducers, dissimilatory sulfite reductase (DsrAB). Results from in vitro sulfite reduction experiments allow us to calculate the in vitro DsrAB isotope effect in 34S/32S (hereafter, ) to be 15.3 ± 2‰, 2σ. The accompanying minor isotope effect in 33S, described as , is calculated to be 0.5150 ± 0.0012, 2σ. These observations facilitate a rigorous evaluation of the isotopic fractionation associated with the dissimilatory MSR pathway, as well as of the environmental variables that govern the overall magnitude of fractionation by natural communities of sulfate reducers. The isotope effect induced by DsrAB upon sulfite reduction is a factor of 0.3–0.6 times prior indirect estimates, which have ranged from 25 to 53‰ in 34εDsrAB. The minor isotope fractionation observed from DsrAB is consistent with a kinetic or equilibrium effect. Our in vitro constraints on the magnitude of is similar to the median value of experimental observations compiled from all known published work, where 34εr−p = 16.1‰ (r–p indicates reactant vs. product, n = 648). This value closely matches those of MSR operating at high sulfate reduction rates in both laboratory chemostat experiments ( 17.3 ± 1.5‰, 2σ) and in modern marine sediments ( 17.3 ± 3.8‰). Targeting the direct isotopic consequences of a specific enzymatic processes is a fundamental step toward a biochemical foundation for reinterpreting the biogeochemical and geobiological sulfur isotope records in modern and ancient environments.

Highlights

Microbial sulfate reduction provides a critical link between Earth’s surface sulfur, carbon, iron, and oxygen cycles (Thode et al, 1961; Holland, 1973; Garrels and Lerman, 1981; Canfield, 2001a)
Key considerations in experimental design are: (i) to provide enough sulfur at each time point for isotopic characterization of residual reactant and products; (ii) to provide the proper reaction conditions to allow for optimal DsrAB activity; (iii) to ensure hydrogenase activity is not inhibited by the experimental pH; and (iv) to ensure the sulfite to hydrogen ratio strongly favors sulfite reduction
In experiments with the D. vulgaris DsrAB, the products were generated with a mean of 19% of the product sulfur forming thiosulfate and the remainder accumulating as trithionate (Figure 2B)

Summary

Introduction

Microbial sulfate reduction provides a critical link between Earth’s surface sulfur, carbon, iron, and oxygen cycles (Thode et al, 1961; Holland, 1973; Garrels and Lerman, 1981; Canfield, 2001a) This metabolism is comprised of a set of enzymes working in concert to reduce sulfate (MSOSR24−g)etnoersautlefisde34(SH/322SS),. Sulfate is first imported into the cytoplasm by a variety of transporters (Cypionka, 1994; Piłsyk and Paszewski, 2009; Figure 1), and subsequently activated to a high-energy intermediate, adenosine 5′-phosphosulfate (APS). The latter reaction generates pyrophosphate (PPi) at the expense of ATP by the enzyme sulfate-adenylyl transferase (Sat) (Peck, 1962). In instances when DsrC is unavailable (e.g., when DsrAB is pure in vitro) or limiting (e.g., intracellular sulfite is in excess of reduced DsrC), intermediates such as thiosulfate (S2O32−) may become important, likely due to the reaction of sulfite with sulfide (in vivo) or the partially reduced sulfur from DsrAB (in vitro)

Objectives

Methods

Results

Discussion

Conclusion