Sulfur biogeochemistry

1. Sulfur biogeochemistry • 8 e- between stable redox states • Polymerizes, cyclizes • Reduced, intermediate, and oxidized solid forms • Thousands of organic sulfur forms (organosulfur compounds; thiols have an –SH group, thioethers –C-S-C, thioesters and sulfonates are oxidized S forms, sulfoxides/sulfones RS(=O)R’, RS(=O)2R, thioketones, thioamides, sulfonium ylides less common)

2. Sulfur Cycle

3. Early earth ocean-atmosphere and S

4. Assimilatory vs. Dissimilatory • S is an essential nutrient (key to amino acids cysteine and methionine) and many other cellular molecules, so all organisms need an assimilatory pathway • Many dissimilatory reactions due to complicated intermedaite pathways involving S redox chemistry- leads to idea that S-utilizing organisms are the most diverse group of microbes which metabolize a single element

5. 1 piece of sulfur oxidation pathways

6. Assimilatory pathways • APS pathway – uptake of SO42- to APS (Adenosine phosphosulfate) using an ATP • APS then goes thorugh 1 of 2 paths: • Forms PAPS (phosphoadenosine phosphosulfate) • Or forms organic thiosulfate derivative (G-S-S-O3) • These are furthur reduced to HS- to form cysteine or other useful sulfur forms • All of this COSTS ENERGY!

7. Dissimilatory SO42- reduction • Biological Sulfate Reduction (BSR) and Thermochemical Sulfate Reduction (TSR) • At temperatures <150-200ºC the reduction of SO42- by reduced organics is VERY slow (though thermodynamically favorable) – formation of sulfide at low T is thus MICROBIAL • ‘Mineralization’ process because H2S and metals strongly interact – form sulfide minerals – very low solubility!

8. Measuring rates of BSR • Profiles and flux rates from gradients • Culture-based incubations • Radiolabeling using 35S-labeld sulfate • Done quickly in sediments (reduce chance of re-oxidation) • Recovery of H2S produced can be difficult (if it quickly goes into pyrite for example it is harder to recover) • However, this is the most accurate and common technique

9. BSR and Carbon mineralization • Carbon compound degradation to CO2 through BSR • AT high sedimentation rates, BSR can account for significant fraction of this • At lower sedimentation rates, BSR is less important • WHY THE DIFFERENCE?? • In lake sediments this can be very different than in marine sediments, WHY?

10. Where do sulfate-reducing bacteria (SRB) hang out? • Need anaerobic/microaerophilic environment, enough SO42-, organics/ H2 • Reduced sediments • Hydrothermal springs (deep sea, terrestrial) • Cyanobacterial mats (where in the mats do you think??) • SRB inhabit widest range of conditions – T 0-127, 0-28% NaCl

11. SRB Phylogeny • Deep-branching, widely distributed across tree of life (both archaeal and bacterial), thermophilic • Bacteria – mostly in d-proteobacteria, also spore-formers, gram+, in nitrospira group • Archaea – Archeoglobus T max=92ºC • LGT of dissimilatory sulfur reductase (DSR) gene supported across archaea, different bacterial species

12. SRB Metabolism pathway • SO42- import – costs energy, coupled to transport of H+ of Na+ • ‘Activated’ by ATP sulfurylase  forms APS, which is then reduced to sulfite which is reduced to sulfide by the DSR enzyme (a reductase) • H2S is highly toxic (interacts strongly with organics and metals)  rapidly excreted from the cell

14. DSR substrate limitations • Require smaller, less recalcitrant substrates (anaerobes do not make radicals needed to degrade bigger molecules into something useable) • Grow best on simple substrates like acetate, but can grow on a wide range of substrates, including some xenobiotics and even PO33- • Some are complete oxidizers, many incomplete – (incomplete ones grow faster) • H2 as an e- source, most are chemolithoheterotrophic, a few known chemolithoautotrophs…

15. SRB Diversity • Over 100 different species known • IN one study, 20 different species were identified from a single sediment sample! • For the same metabolism – what other factors may play into which one(s) are predominant at any point in time or space??

16. Elemental sulfur • S8 a product of sulfide oxidation, some organisms store it intracellualry, also forms abiotically on interaction of H2S with metals, organics • Elemental sulfur respiration coupled with H2 or organic carbon oxidation (complete and incomplete) found in many organisms • Several identified species of the d-proteobacterial clade that primarily metabolize S8, • Widespread archaeal metabolism – Crenarcheota, Sulfolobus, Acidianus, othrs

17. Sulfide oxidation • Abiotic pathways – sulfide reaction with FeOOH or MnOOH is fast, reaction with O2 slower, with NO3- slow too… • Plenty of differences in the intermediates of H2S oxidation depending on specific chemistry and availability of oxidants too

18. Black Sea

19. Green Lake, NY • Voltammetric evidence for significant role of polysulfides in sulfide oxidation and elemental sulfur reactions

20. Sulfide Oxidizing Organisms • Chemolithoautotrophs (and heterotrophs) exist that can oxidize H2S and other intermediates • Many can also reduce elemental sulfur… • Use O2 or NO3- as electron acceptor • Most obligate or facultative aerobes, but some are obligately microaerophilic (can’t handle above a few tens of uM)

21. Intracellular S8 • Several S-oxidizers can store S8 in vacuoles • Noteably Beggiatoa and Thiothrix spp.

22. HS- Sxn- S8 S4O62- HSO3- S2O32- Cave formation and stratified analogues in central Italy • Influx of sulfide-rich water accelerates cave formation: H2S + 2 O2 SO42- + 2 H+ CaCO3 + H+ Ca2+ + HCO3-

23. 3 different predominant mat types Current (mA) Potential (V vs. Ag/AgCl) Microbial ecology and sulfur speciation • Different microbial communities found in different places --- related to BIG changes in S speciation! Sxn- White: d-proteobacterial mat Red: thiovulum mat Green: beggiotoa mat HS- S8 HSO3- S2O32-

24. thiovulum mats Above (green) and into biofilm (others) HS- Sxn- S8 Current (mA) S2O32- Potential (V vs. Ag/AgCl) Scans above (green and into biofilm, red) HS- S8 beggiatoa mats Current (mA) Potential (V vs. Ag/AgCl) ‘d-proteobacterial’ mats Scans into white mat material Sxn- HS- S8 Current (mA) Potential (V vs. Ag/AgCl)

25. ‘thiovulum’ mats, Pozzo di Cristale, Frassassi caves

26. Current (mA) Potential (V vs. Ag/AgCl) Thiovulum mat profile data ~ 50 mm thick biofilm above

27. Current (mA) Potential (V) vs Ag/AgCl Snottite electrochemistry pH varies 1-3 in these snottite streamers

28. S8 in biofilms at Frasassi Images courtesy Jenn Macalady, Penn State

29. 16s library of the biofilms in Frassassi • New results looking at metagenomic data has identified a gene regulating elemental sulfur ‘docking’ Courtesy Macalady lab, Penn State

30. Phototrophic S-oxidation • Anoxygenic phototrophy using H2S, S8, S2O32- as electron donors • Organisms are common, in 5 major groups: • Purple sulfur bacteria • Purple nonsulfur bacteria • Green sulfur bacteria • Green nonsulfur bacteria • Heliobacteria • These archaic groupings derived from ‘sulfur’ groups depositing visible S8, nonsulfur ones did not – mistakenly thought they did not use reduced sulfur as a result, and we still use the names…

31. Phototrophic Mats - Cyanos Anoxygenic photosynthetic organisms oxidizing H2S across a VERY sharp gradient!! Electrode tip stuck bottom

32. Phototrophic mats - PSB • Purple sulfur bacteria mats • Respond to light level changes in minutes  position in sediment and water column can vary significantly!

33. Light Manipulation experiments Jacket on Jacket off Hat on Hat off

34. S-oxidizer phylogeny • Anoxygenic photosynthesis development before oxygenic photosynthesis? • Geochemical record of the earth’s oceans? • Photosystem less complicated • Anoxygenic organisms more deeply branching • Others argued based on pigment biosynthesis pathways oxygenic photosynthesis is first • Subsequent genetic analysis using genes related to pigment biosynthesis showed anoxygenic photosynthesis first (specifically, PSB) – but here are some complications involving possible LGT…

35. Disproportionation • Sulfur’s equivalence to fermentation – intermediate oxidation state sulfur species (elemental sulfur, thiosulfate, sulfite) split into one more and one less oxidized forms, ex: • S2O32- + H2O  H2S + SO42-

36. S stable isotopes • 4 stable isotopes of sulfur: 32S (95.04%), 33S (0.749%), 34S (4.20%), 36S (0.0156%) • Thermodynamic equilibrium for the fractionation of S isotopes rarely obtained – observed fractionations largely kinetic • SRB fractionations (cultures) 3-46‰ • Rates, species/enzymes, substrates affect this • S-disproportionation also results in large fractionation (up to 37‰) • SRB fractionations in nature up to >100+‰ • S-oxidation (biotic or abiotic) does not produce much fractionation at all!