Molybdenum
1900 – discovered in plant ashes
1930 - azotobacter (N2 fixer) growth requirement for Mo
1940 – established as essential micronutrient for plants
1950 - [Mo] affected rat XO activity; [Mo]~ [flavin]
Mo associated with aldehyde oxidase in rabbit livers
1960 - Mo crucial to N-cycle
Mo

The only 4d metal essential to Life
S
Table 1. The Oxomolybdenum Enzymes
The Xanthine Oxidase Family (LMoOS-Possessing Enzymes)
enzyme
source
subunits
cofactora
xanthine oxidase cow’s milkb R2 MPT
xanthine dehydrogenase chicken liverc R2 MPT
rat liverd R2 MPT
Micrococcus lactyliticuse
Drosophila melanogasterf R2 MPT
Chlamydomonas reinhardtiig
humanh R2 (MPT)
aldehyde oxidase rabbit liveri R2 (MPT)
humanj R2 (MPT)
cowk R2 (MPT)
aldehyde oxidoreductase (dehydrogenase) Desulfovibrio gigasl R2 MCD
Acetobacter polyoxogenesm
formate dehydrogenase Alcaligenes eutrophusn Râçä
Methylosinus trichosporumo R2â2ç2ä2
CO dehydrogenase (oxidoreductase) Pseudomonas carboxydovoransp R2â2ç2
Pseudomonas carboxydoflavaq R2â2ç2 MCD
Oligotropha carboxidovoransr R2â2ç2 MCD
quinoline-2-oxidoreductase Pseudomonas putida s R2â2ç2 MCD
Rhodococcus sp. B1t R2â2ç2 MCD
Comamonas testosteroni 63u R2â2ç2 MCD
isoquinoline 1-oxidoreductase Pseudomonas diminutav Râ MCD
quinoline-4-carboxylate-2-oxidoreductase Agrobacterium sp. 1Bw R2â2ç2 MCD
quinaldine-4-oxidoreductase Arthrobacter sp.x R2â2ç2 MCD
quinaldic acid 4-oxidoreductase Serratia marcescensy
Pseudomonas sp. AK-2z Râ
nicotinic acid hydroxylase (dehydrogenase) Clostridium barkeriaa,ab R2
Bacillus niaciniac R2â2ç2
Arthrobacter oxidansad Râç
6-hydroxynicotinate hydroxylase Bacillus niaciniac Râç
nicotine dehydrogenase Arthrobacter oxidansad Râç
Arthrobacter nicotinovoransae
picolinate hydroxylase Arthrobacter picolinophilusaf R2â2ç2 MCD
(2R)-hydroxycarboxylate oxidoreductase Proteus vulgarisag
(R. Hille, Chem. Rev.
1996)
The Sulfite Oxidase Family (LMoO2-Possessing Enzymes)
sulfite oxidase bovine liverah R2 MPT
chicken liverai R2 MPT
rat liveraj R2 MPT
humanak R2 MPT
Thiobacillus novellusal R
nitrate reductase (assimilatory) Neurospora crassaam R2 MPT
spinachan R2 MPT
Chlorella vulgarisao R4 MPT
The DMSO Reductase Family (L2MoX-Possessing Enzymes)
DMSO reductase Rhodobacter sphaeroidesap R MGD
Rhodobacter capsulatus aq R MGD
Escherichia coli ar Râç MGD
biotin-S-oxide reductase Escherichia colias
trimethylamine-N-oxide reductase Escherichia coliat R2
nitrate reductase (dissimilatory) Escherichia coli (NarGHI)au Râç MGD
Escherichia coli (NarZYV)av Râç MGD
Escherichia coli (FdoGHI)aw Râç MGD
Paracoccus denitrificans (NapABCD)ax Râçä
Haloferax volcaniiay
formate dehydrogenase Escherichia coli (FdhF)az R MGD
Escherichia coli (FdnGHI)ba Râç MGD
Escherichia coli (FdoGHI)bb Râç MGD
Methanobacterium formicicumbc (XdS) Râç MGD
Wollinella succinogenesbd Râç
polysulfide reductase Wolinella succinogenesbe Râç MGD
arsenite oxidase Alcaligenes faecalisbf R MCD
formylmethanofuran dehydrogenase) Râçä(ú) MGD, MAD, MHD
Methanosarcina barkeribh MGD
Unclassified Molybdenum-Containing Enzymes
pyridoxal oxidase Drosophila melanogaster
chlorate reductase Proteus mirabilis
tetrathionite reductase Proteus mirabilis
xanthine dehydrogenase Clostridium sp
pyruvate:ferredoxin oxidoreductase Archaeoglobus fulgidus
pyrogallol transhydroxylase Pelobacter acidigallici
2-furoyl-CoA dehydrogenase Pseudomonas putida
Locating the Molybdenum Cofactor
All plants require the
molybdenum enzyme
All mammals require
molybdenum enzymes
Nitrate Reductase
Sulfite Oxidase
NO3- + 2H+ + 2e-
SO32- + H2O
NO2- + H2O
SO42- + 2H+ + 2e-
Mo(4+) + X-O + 2H+
Mo(6+) + X + H2O
NO2-
Nitrate
reductase
N2
nitrogenase
Nitrogen- cycle
NO3-
NH3
polysulfide
reductase
SH2
CH4
Sx
Carbon- cycle
Sulfur- cycle
CO2
CH3CO2-
(CH3)2S
(CH3)2SO
DMSO
reductase
Table 1. The Oxomolybdenum Enzymes
(R. Hille, Chem. Rev. 1996)
The Xanthine Oxidase Family (LMoOS-type)
enzyme
xanthine oxidase
xanthine dehydrogenase
source
cofactor
cow’s milk
MPT
chicken liver
MPT
rat liver
MPT
Drosophila melanogaster
MPT
human
MPT)
aldehyde oxidase
rabbit liver
MPT
human
MPT
cow
MPT
aldehyde oxidoreductase (dehydrogenase) Desulfovibrio gigas MCD
formate dehydrogenase
Alcaligenes eutrophus
CO dehydrogenase (oxidoreductase) Pseudomonas carboxydovorans
quinoline-2-oxidoreductase
Pseudomonas putida
MCD
isoquinoline 1-oxidoreductase Pseudomonas diminuta
MCD
quinoline-4-carboxylate-2-oxidoreductase Agrobacterium sp
MCD
quinaldine-4-oxidoreductase
Arthrobacter sp.
MCD
quinaldic acid 4-oxidoreductase Serratia marcescens
Pseudomonas sp. AK
nicotinic acid hydroxylase (dehydrogenase) Clostridium barkeriaa,
6-hydroxynicotinate hydroxylase Bacillus niaciniac
nicotine dehydrogenase
Arthrobacter oxidansa
The Sulfite Oxidase Family (LMoO2-type)
enzyme
source
sulfite oxidase
bovine liver
chicken liver
rat liver
human
Thiobacillus novellus
nitrate reductase (assimilatory) Neurospora crassa
spinach
Chlorella vulgari
The DMSO Reductase Family (L2MoX-type)
DMSO reductase
Rhodobacter sphaeroides
biotin-S-oxide reductase
Escherichia colia
trimethylamine-N-oxide reductase Escherichia colia
cofactor
MPT
MPT
MPT
MPT
MPT
MPT
MPT
MGD
nitrate reductase (dissimilatory) Escherichia coli (NarGHI)
MGD
formate dehydrogenase
Escherichia coli (FdhF)
MGD
polysulfide reductase
Wolinella succinogene
MGD
arsenite oxidase
Alcaligenes faecalisb
MCD
formylmethanofuran dehydrogenase)
MGD, MAD, MHD
Unclassified Molybdenum-Containing Enzymes
pyridoxal oxidase
Drosophila melanogaster
chlorate reductase
Proteus mirabilis
tetrathionite reductase
Proteus mirabilis
xanthine dehydrogenase
Clostridium sp.
pyruvate:ferredoxin oxidoreductase Archaeoglobus fulgidus
pyrogallol transhydroxylase
Pelobacter acidigallici
2-furoyl-CoA dehydrogenase Pseudomonas putida
The Generic Molybdenum Cofactor
MAD
MGD
MCD
MPT
There’s not only one, but a family of Moco’s
Why molybdopterin?
dithiolene
Mo
pterin
Table 1. The Oxomolybdenum Enzymes
(R. Hille, Chem. Rev. 1996)
The Xanthine Oxidase Family (LMoOS-type)
The Sulfite Oxidase Family (LMoO2-type)
The DMSO Reductase Family (L2MoX-type)
Used to be classed with XO
b/c structural similarity.
Unique Mo enzyme with a 2nd metal, Cu
Table 1. The Oxomolybdenum Enzymes
(R. Hille, Chem. Rev. 1996)
The Xanthine Oxidase Family (LMoOS-type)
The Sulfite Oxidase Family (LMoO2-type)
Xanthine Oxidase/Dehydrogenase
- in mammalian milk but >50% de-Mo
- in liver: purine metabolism (gout)
- implicated in heart damage (ROS form’n)
Nitrate Reductase
- in all plants
- assimilatory form (plants, fungi, algae)
The DMSO Reductase Family (L2MoX-type)
- all bacterial or from archea
- many in respiration using specific e- acceptors
- recall DMSOR in cloud formation ….
- simplest Mo-enzymes: w/o other cofactors
Sulfite Oxidase
- most critical for humans
- detoxification of sulfite
- also has role in S-metabolism,
from Cys  Met   SO32-  SO42-
Table 1. The Oxomolybdenum Enzymes
(R. Hille, Chem. Rev. 1996)
The Xanthine Oxidase Family (LMoOS-type)
- in mammalian milk but >50% de-Mo
- in liver: purine metabolism (gout)
- implicated in heart damage (ROS form’n)
The Sulfite Oxidase Family (LMoO2-type)
Nitrate Reductase
- in all plants
- assimilatory form (plants, fungi, algae)
NO3- + 2H+ + 2e-  NO2- + H2O
The DMSO Reductase Family (L2MoX-type)
- all bacterial or from archea
- many in respiration using specific e- acceptors
- recall DMSOR in cloud formation ….
- simplest Mo-enzymes: w/o other cofactors
Sulfite Oxidase
- most critical for humans
- detoxification of sulfite
- also has role in S-metabolism,
from Cys  Met   SO32-  SO42SO32- + H2O  SO42- + 2H+ + 2e-
Root nodules
formed
by
Rhizobium bacteria
living symbiotically
through
nitrogen fixation.
Tomato plants with and without molybdenum available.
Tobacco plants (Arabidopsis thaliana)
Nitrate
Nitrite
Proteins
nitrate
Healthy
(wild type)
All plants require
Assimilatory
Nitrate Reductase
Sick
(mutant)
nitrite
Introduction
Repairing the Molybdenum Cofactor
Baby Z Cured of Rare Disease in 3 Days
University of Arizona, Tucson, October 2010
(Southern Health/AFP/Getty Images)
Orphan Drug Treatment Used Only on
Mice to Get Hearing Before FDA
Baby Z had a one in a million
By SUSAN
chance of developing
rare9, 2009
DONALDSON
JAMES, aNov.
metabolic disorder called
molybdenum cofactor deficiency
and zero chance of avoiding the
inevitable death sentence that
comes with it.
Australian
girl had acofactor,
Worldwide, there are only about 50The
cases
of molybdenum
seemingly
normal
in May
or sulfite oxidase deficiency, mostly
in Europe
and inbirth
the United
2008 but,ofwithin
hours, she began
States, according to the National Institutes
Health.
havingismultiple
as
Molybdenum, like other organic metals,
essentialseizures
for the -human
many asmolecule
10 an hour
as sulfite
body. Its cofactor is a small, complicated
that--acts
as a
Why the correct oxidation state matters
MRI of brain of deceased baby
with Sulfite Oxidase Deficiency
MRI of healthy brain
The baby died because this reaction didn’t happen:
SO32- +
Sulfite
S4+
H2O
--->
SO42- + 2H+ +
Sulfate
S6+
2e-
The baby has a genetic defect in the enzyme
that catalyzes this reaction.
Babies with this genetic disease die within hours.
The enzyme is Sulfite Oxidase.
You have it in your liver.
Review the terms:
Oxidation
half reaction
Reduction
half reaction
SO32- + H2O
S4+ in Sulfite
--->
SO42- + 2H+ + 2eS6+ in Sulfate
Mo6+ + 2e- ---> Mo4+
Net redox reaction
SO32- + H2O + Mo6+ --->
SO42- +
2H+ + Mo4+
Is this reaction spontaneous?
Caroline Kisker
Würzburg, Germany
Protein crystallographer
X-ray structure of chicken liver Sulfite Oxidase
Kisker, Enemark
Moco locked into position by H-bonds to pterin
A catalytic cycle for how Mo oxidizes SO32coupled
oxygen
electron
proton
transfer
S
atom
+ SO3 2-
O
transfer
+6
Mo
S
O
O
S
O
- H+, - e-
O
S
O
O
Mo+5
OH
O
- H+, - e- SO42-
S
S
O
O
S
S
a
S
Mo
S
S
O
S
Mo+4
O H
H
S
+ H2O
Mo+4
O O
Mo+4 S
O
O
Water mediated OAT
Similar cycles can be devised for
Sulfite Oxidase
and Nitrate Reductase
SO32- + H2O
Sulfite
S4+
NO3- +
Nitrate
N5+
---> SO42- + 2H+ +
Sulfate
S6+
2H+ +
2e-
2e- ---> NO2- + H2O
Nitrite
N3+
Environmental impact of DMSOR and the smell of the ocean
DMS
CH3SO3- cloud nucleation sites
hn, photo-oxidation
DMS
(CH3)2S(CH2)2CO2from algae
DMSO Reductase
DMSO
DMSOR Structure: Controversy #2
The first Mo enzyme X-ray structure: DMSO Reductase
Group Meeting Bryn Mawr College, October 2010
Doug Rees, 1996
Doug Rees, Ca
Protein crystal
SURPRISE!!!!
• 2 molydopterin ligands!
• nucleoside termini on pterin
• very long Mo-S bonds
DMSOR Structure: Controversy #2
The first look at molybdopterin was on a tungsten enzyme!
Hyperthermophilic TungstonEnzyme, Aldehyde Ferredoxin Oxidoreductase
Group Meeting Bryn Mawr College, October 2010
Doug Rees et al., Science,1995
SURPRISE!!!!
• not the molydopterin ligand!
• is that pyran ring actually right???
DMSOR Structure: Controversy #2
Group Meeting Bryn Mawr College, October 2010
Will the real active site structure in DMSO Reductase please stand up?
(S J N Burgmayer, in Progress in Inorganic Chemistry, 2004)
And the answer was:
Hermann Schindelin, Würzburg, Germany
Protein crystallographer
DMSOR Structure: Controversy #2
Group Meeting Bryn Mawr College, October 2010
1.3 Å X-ray Structure in DMSO Reductase (Schindelin)
Active form
What does it mean?
There are 2
superimposed
structures.
(only one is inactive!)
Inactive
form