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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)
Sulfur Cycle
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
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!
Dissimilatory SO42- reduction
• Biological Sulfate Reduction (BSR) and Thermochemical Sulfate Reduction (TSR)
• At temperatures <150-200ºC the reduction of SO4
2- 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!
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
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?
Where do sulfate-reducing bacteria (SRB) hang out?
• Need anaerobic/microaerophilic environment, enough SO4
2-, 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
SRB Phylogeny• Deep-branching, widely distributed across
tree of life (both archaeal and bacterial), thermophilic
• Bacteria – mostly in -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
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
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 PO3
3-
• Some are complete oxidizers, many incomplete – (incomplete ones grow faster)
• H2 as an e- source, most are chemolithoheterotrophic, a few known chemolithoautotrophs…
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??
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 -proteobacterial clade that primarily metabolize S8,
• Widespread archaeal metabolism – Crenarcheota, Sulfolobus, Acidianus, othrs
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
Green Lake, NY
• Voltammetric evidence for significant role of polysulfides in sulfide oxidation and elemental sulfur reactions
Green Lake Voltammetric Profile
0
5
10
15
20
25
0 0.1 0.2 0.3Peak Current (A)
Dep
th(m
)
Oxygen(dissolved)
HydrogenSulfide
Polysulfide
ElementalSulfur
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)
Intracellular S8
• Several S-oxidizers can store S8 in vacuoles
• Noteably Beggiatoa and Thiothrix spp.
Cave formation and stratified analogues in central Italy
• Influx of sulfide-rich water accelerates cave formation: H2S + 2 O2 SO4
2- + 2 H+
CaCO3 + H+ Ca2+ + HCO3-
1.375
-0.230
0.000
0.200
0.400
0.600
0.800
1.000
1.200
-0.050-1.800 -1.500 -1.250 -1.000 -0.750 -0.500 -0.250
S8
Sxn- HS-
S2O32-
HSO3-
S4O62-
Microbial ecology and sulfur speciation• Different microbial communities found in different
places --- related to BIG changes in S speciation!
3 different predominant mat types0.871
-0.265
-0.200
-0.100
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0.800
-0.050-1.800 -1.500 -1.250 -1.000 -0.750 -0.500 -0.250
Potential (V vs. Ag/AgCl)
Cur
rent
(A
)
White: -proteobacterial matRed: thiovulum matGreen: beggiotoa mat
S8
Sxn-
HS-
S2O32-HSO3
-
‘-proteobacterial’ mats 0.997
-0.331
-0.200
-0.100
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0.800
0.900
-0.050-1.800 -1.500 -1.250 -1.000 -0.750 -0.500 -0.250
Potential (V vs. Ag/AgCl)
Cur
rent
(A
)
Scans into white mat material
Sxn-
Potential (V vs. Ag/AgCl)
Cur
rent
(A
)
1.275
-0.294
-0.200
0.000
0.200
0.400
0.600
0.800
1.000
1.200
-0.050-1.800 -1.600 -1.400 -1.200 -1.000 -0.800 -0.600 -0.400 -0.200
Scans above (green and into biofilm, red)
Potential (V vs. Ag/AgCl)
Cur
rent
(A
)
0.420
-0.107
-0.050
0.000
0.050
0.100
0.150
0.200
0.250
0.300
0.350
-0.050-1.800 -1.500 -1.250 -1.000 -0.750 -0.500 -0.250
Above (green) and into biofilm (others)
beggiatoamats
thiovulummats
S8
S8
S8
Sxn-
HS-
HS-
HS-
S2O32-
Thiovulum mat profile data
~ 50 m thick biofilm
Thiovulum Mat Profile Pozzo di Cristale
0
10
20
30
40
50
60
70
80
90
100
0 20 40 60
Current (nA)
De
pth
fro
m b
otto
m (
um
)
elemental sulfur
sulfite
sulfide (uM)
Potential (V vs. Ag/AgCl)
Cur
rent
(A
)
0.420
-0.107
-0.050
0.000
0.050
0.100
0.150
0.200
0.250
0.300
0.350
-0.050-1.800 -1.500 -1.250 -1.000 -0.750 -0.500 -0.250
above
Snottite electrochemistry
Potential (V) vs Ag/AgCl
Cu
rren
t (
A)
0.278
-0.108
-0.075
-0.050
-0.025
0.000
0.025
0.050
0.075
0.100
0.125
0.150
0.175
0.200
0.225
0.250
-0.050-1.300 -1.000 -0.800 -0.600 -0.400 -0.200
Sulfide peak location on Au/Hg microelectrode in Mini-Primrose water over a range of pH values on HMDE
y = -0.0702x - 0.2255
R2 = 0.9837
-0.7
-0.65
-0.6
-0.55
-0.5
-0.45
-0.4
-0.35
-0.3
1 2 3 4 5 6 7
pH
Po
ten
tial
(V
vs.
Ag
/Ag
Cl)
pH varies 1-3 in these snottite streamers
Courtesy Macalady lab, Penn State
16s library of the biofilms in Frassassi
• New results looking at metagenomic data has identified a gene regulating elemental sulfur ‘docking’
Phototrophic S-oxidation• Anoxygenic phototrophy using H2S, S8, S2O3
2- 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…
Phototrophic Mats - CyanosPhototrophic Mat outside fracture
spring - Frassassi
-350
-300
-250
-200
-150
-100
-50
0
0 20 40 60
Conc (nA)
Dep
th (
m)
sulfide (uM)
elementalsulfur
oxygen
approximatetop of mat
Anoxygenic photosynthetic organisms oxidizing H2S across a VERY sharp gradient!!
Electrode tip stuck bottom
Phototrophic mats - PSB• Purple sulfur bacteria mats
– Respond to light level changes in minutes position in sediment and water column can vary significantly!
Purple sulfur bacteria mats
-800
-700
-600
-500
-400
-300
-200
-100
0
0 500 1000 1500 2000
H2S(aq) Concentration (M)
Dep
th (
mic
ron
s)
Light Manipulation experimentsCyanobacteria Light Manipulation Experiment
0
50
100
150
200
250
300
350
400
450
-80 -60 -40 -20 0 20 40 60 80 100 120
time (seconds)
nA
H2S
Jacket on Jacket off Hat on Hat off
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…
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:– S2O3
2- + H2O H2S + SO42-
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!