Information on EC 4.99.1.3 - sirohydrochlorin cobaltochelatase

Word Map on EC 4.99.1.3
Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
Specify your search results
Select one or more organisms in this record:
Show additional data
Do not include text mining results
Include (text mining) results (more...)
Include results (AMENDA + additional results, but less precise; more...)


The expected taxonomic range for this enzyme is: Archaea, Bacteria

EC NUMBER
COMMENTARY hide
4.99.1.3
-
RECOMMENDED NAME
GeneOntology No.
sirohydrochlorin cobaltochelatase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
cobalt-sirohydrochlorin + 2 H+ = sirohydrochlorin + Co2+
show the reaction diagram
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
cob(II)yrinate a,c-diamide biosynthesis I (early cobalt insertion)
-
-
Metabolic pathways
-
-
Porphyrin and chlorophyll metabolism
-
-
vitamin B12 metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
cobalt-sirohydrochlorin cobalt-lyase (sirohydrochlorin-forming)
This enzyme is a type II chelatase, being either a monomer (CbiX) or a homodimer (CibK) and being ATP-independent. CbiK from Salmonella enterica uses precorrin-2 as the substrate to yield cobalt-precorrin-2. The enzyme contains two histidines at the active site that are thought to be involved in the deprotonation of the tetrapyrrole substrate as well as in metal binding. CbiX from Bacillus megaterium inserts cobalt at the level of sirohydrochlorin (factor-II) rather than precorrin-2.
CAS REGISTRY NUMBER
COMMENTARY hide
81295-49-0
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain DSM509
-
-
Manually annotated by BRENDA team
genes cobN, cobS and cobT encoding the tree subunits
-
-
Manually annotated by BRENDA team
Methanobacter thermoautotrophicum
-
-
-
Manually annotated by BRENDA team
strain Fusaro
-
-
Manually annotated by BRENDA team
no activity in Bacillus subtilis
-
-
-
Manually annotated by BRENDA team
no activity in eukaryota
-
-
-
Manually annotated by BRENDA team
strain PCC6803
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
-
a cbiX-deficient mutant is unable to respire anaerobically on nitrate
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + sirohydrochlorin + Co2+
ADP + phosphate + Co(II)-sirohydrochlorin + H+
show the reaction diagram
ATP + sirohydrochlorin + Fe2+
ADP + phosphate + Fe(II)-sirohydrochlorin + H+
show the reaction diagram
cobalt-sirohydrochlorin + H+
sirohydrochlorin + Co2+
show the reaction diagram
precorrin-2 + Co2+
Co-precorrin-2 + H+
show the reaction diagram
precorrin-2 + Co2+
cobalt-precorrin-2 + H+
show the reaction diagram
-
-
-
?
sirohydrochlorin + Co2+
cobalt-sirohydrochlorin + 2 H+
show the reaction diagram
sirohydrochlorin + Co2+
cobalt-sirohydrochlorin + H+
show the reaction diagram
sirohydrochlorin + Co2+
cobaltsirohydrochlorin + H+
show the reaction diagram
sirohydrochlorin + Fe2+
iron-sirohydrochlorin + 2 H+
show the reaction diagram
sirohydrochlorin + Fe2+
siroheme + H+
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
cobalt-sirohydrochlorin + H+
sirohydrochlorin + Co2+
show the reaction diagram
precorrin-2 + Co2+
Co-precorrin-2 + H+
show the reaction diagram
-
tetrapyrrole biosynthesis, formation of tetrapyrrole cofactors
-
?
sirohydrochlorin + Co2+
cobalt-sirohydrochlorin + 2 H+
show the reaction diagram
sirohydrochlorin + Co2+
cobalt-sirohydrochlorin + H+
show the reaction diagram
sirohydrochlorin + Co2+
cobaltsirohydrochlorin + H+
show the reaction diagram
sirohydrochlorin + Fe2+
iron-sirohydrochlorin + 2 H+
show the reaction diagram
Q72CB8, Q72EC8
-
-
-
?
sirohydrochlorin + Fe2+
siroheme + H+
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ni2+
Methanobacter thermoautotrophicum
-
able to chelate nickel
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00043 - 0.00062
sirohydrochlorin
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0015
purified recombinant CbiKc, with Fe2+
0.0056
His-tagged protein of mutant M508
0.007
His-tagged protein of mutant M518
0.0075
His-tagged protein of mutant M54
0.01098
His-tagged protein of mutant M136
0.0135
His-tagged protein of mutant M507
0.0155
His-tagged protein of mutant M519
0.0164
His-tagged protein of mutant M10
0.017
His-tagged protein
0.018
Methanobacter thermoautotrophicum
-
-
0.0255
His-tagged protein of mutant M510
0.0272
His-tagged protein of mutant M51
0.06
Methanobacter thermoautotrophicum
-
specific activity to chelate nickel
0.08
-
overproduced in Escherichia coli
0.0938
His-tagged protein of mutant M200
0.1197
His-tagged wild type protein
1.4
-
CbiX6H, overproduced in Escherichia coli
1.5
substrate Fe2+, pH 8.0
4
substrate Co2+, pH 8.0
13
substrate Fe2+, pH 8.0
22
substrate Co2+, pH 8.0
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Archaeoglobus fulgidus (strain ATCC 49558 / VC-16 / DSM 4304 / JCM 9628 / NBRC 100126)
Archaeoglobus fulgidus (strain ATCC 49558 / VC-16 / DSM 4304 / JCM 9628 / NBRC 100126)
Archaeoglobus fulgidus (strain ATCC 49558 / VC-16 / DSM 4304 / JCM 9628 / NBRC 100126)
Archaeoglobus fulgidus (strain ATCC 49558 / VC-16 / DSM 4304 / JCM 9628 / NBRC 100126)
Desulfovibrio vulgaris (strain Hildenborough / ATCC 29579 / DSM 644 / NCIMB 8303)
Desulfovibrio vulgaris (strain Hildenborough / ATCC 29579 / DSM 644 / NCIMB 8303)
Desulfovibrio vulgaris (strain Hildenborough / ATCC 29579 / DSM 644 / NCIMB 8303)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
14300
SDS-PAGE, His-tagged protein of mutant M76
14900
SDS-PAGE, His-tagged wild type protein
15000
-
SDS-PAGE
17000
Methanobacter thermoautotrophicum
-
SDS-PAGE
17200
SDS-PAGE, His-tagged protein of mutant M200
17500
SDS-PAGE, His-tagged protein of mutant M54
17800
SDS-PAGE, His-tagged protein of mutant M136
18400
SDS-PAGE, His-tagged protein of mutant M150
18600
SDS-PAGE, His-tagged protein of mutant M10
18700
SDS-PAGE, His-tagged protein of mutant M508
20100
SDS-PAGE, His-tagged protein of mutant M507
23000
-
x * 23000, SDS-PAGE
25300
SDS-PAGE, His-tagged protein of mutant M518
27000
SDS-PAGE, His-tagged protein of mutant M51
28000
x * 28000, SDS-PAGE and calculated, mature protein
31000
x * 31000, PAGE and calculated, mature protein; x * 31000, recombinant enzyme, SDS-PAGE
35000
-
SDS-PAGE
37000
-
x * 137000, subunit CobN, + x * 37000, subunit CobS, + x * 71000, subunit CobT
38000
-
SDS-PAGE
71000
-
x * 137000, subunit CobN, + x * 37000, subunit CobS, + x * 71000, subunit CobT
137000
-
x * 137000, subunit CobN, + x * 37000, subunit CobS, + x * 71000, subunit CobT
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oligomer
-
x * 137000, subunit CobN, + x * 37000, subunit CobS, + x * 71000, subunit CobT
tetramer
additional information
-
the molecules are arranged in a two-tiered ring structure with the six subunits in each ring organized as a trimer of dimers, subunits CobS and CobT form a chaperone-like complex. Homology modelling of subunit CobS, overview
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
using the sitting drop vapor diffusion technique in 96-well plates, in two different crystal forms consisting of a central mixed b-sheet flanked by four alpha helices
hanging drop vapor diffusion method, using 100 mM Tris-HCl. pH 8.5, containing 2.0 M ammonium sulfate
using the sitting drop vapor diffusion technique in 96-well plates, in two different crystal forms consisting of a central mixed b-sheet flanked by four alpha helices
hexagonal crystals are grown using the hanging-drop method, crystals are in space group P6(3)22 with cell dimensions a = b = 128.08 A, c = 85.44 A
-
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
requires molecular oxygen
-
650801
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
by using a metal chelating affinity column charged with Ni2+ or a chelating Sepharose-column charged with Co2+
by using metal chelate affinity chromatography
Ni-Sepharose column chromatography
-
purified anearobically
-
recombinant His-tagged wild-type isozyme and mutant DELTA28DVU0650 from Escherichia coli by nickel affinity chromatography, dialysis, anion exchange chromatography and gel filtration; recombinant His-tagged wild-type isozyme from Escherichia coli by nickel affinity chromatography, dialysis, anion exchange chromatography and gel filtration
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned and overexpressed in Escherichia coli and Bacillus megaterium
-
expressed in Escherichia coli BL21 codonplus (RIPL) cells
-
expression in Escherichia coli; expression in Escherichia coli; gene DVU0650, DNA and amino acid sequence determination and analysis, expression of wild-type enzyme and mutant DELTA28DVU0650 in Escherichia coli as His-tagged proteins, complementation of Escherichia coli cysG mutant strain by DELTA28CbiKP; gene DVU1365, DNA and amino acid sequence determination and analysis, expression of wild-type enzyme in Escherichia coli as His-tagged protein, complementation of Escherichia coli cysG mutant strain by CbiKC
genes cobN, cobS and cobT, DNA and amino acid sequence determination and analysis, overexpression in Escherichia coli strain BL21(DE3)
-
overexpressed in Escherichia coli
-
overexpressed in Escherichia coli BL21(DE3)pLys cells by cloning the gene into pET14b
-
overexpression in Escherichia coli as non His-tagged protein, as His-tagged protein fusion protein and as selenomethionine-substituted protein
overexpression in siroheme deficient Escherichia coli after random in vitro gene sequence rearrangements in order to screen the generated library for Escherichia coli cells that are able to insert both cobalt and to a lesser extend iron into its tetrapyrrole substrate sirohydrochlorin
overproduced in Escherichia coli
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
10 nM exogenous cobalt decreases the levels of the enzyme 13fold
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C259S
-
site-directed mutagenesis
C262S
-
site-directed mutagenesis
M257L
-
site-directed mutagenesis
C259S
-
site-directed mutagenesis
-
C262S
-
site-directed mutagenesis
-
M257L
-
site-directed mutagenesis
-
H12A
wild type enzyme
H192A
double mutation in M518 mutant; double mutation in M51 mutant
H78A
double mutation in M51 mutant; wild type enzyme
H127A
-
inactive
H187A
-
inactive
H145A/H207A
-
site-directed mutagenesis
additional information
Show AA Sequence (1582 entries)
Please use the Sequence Search for a specific query.