Metabolic Modeling Tutorial
discounted EARLY registration ends Dec 31, 2014
Metabolic Modeling Tutorial
discounted EARLY registration ends Dec 31, 2014
Metabolic Modeling Tutorial
discounted EARLY registration ends Dec 31, 2014
Metabolic Modeling Tutorial
discounted EARLY registration ends Dec 31, 2014
Metabolic Modeling Tutorial
discounted EARLY registration ends Dec 31, 2014
twitter

Escherichia coli K-12 substr. MG1655 Enzyme: molybdenum cofactor guanylyltransferase



Gene: mobA Accession Numbers: EG11829 (EcoCyc), b3857, ECK3849

Synonyms: chlB, mob, narB, molybdopterin-guanine dinucleotide, protein Ar, protein FA

Regulation Summary Diagram: ?

Summary:
MobA is required for the biosynthesis of the molybdopterin guanine dinucleotide (MGD) cofactor [Johnson91]. The enzyme catalyzes the transfer of a GMP moiety from GTP to Mo-molybdopterin, forming MGD [Neumann11]. The MGD cofactor can be further modified to bis-MGD. Formation of bis-MGD is proposed to involve formation of a bis-Mo-MPT intermediate followed by the addition of two GMP moieties by MobA. This cofactor is then transferred to its target protein TorA [Reschke13].

MobA is monomeric in solution [Palmer94]. Crystal structures of MobA have been solved [Lake00, Stevenson00, Guse03]. Site-directed mutagenesis of conserved residues showed that Gly15 is essential for MobA function, and Gly82 stabilizes the product-bound form of the enzyme [Guse03]. The enzyme's specificity for GTP was also investigated [Neumann11].

MobA directly interacts with both MoeA and MobB [Magalon02]. In the presence of the enzyme-specific chaperone NarJ and the mature molybdenum cofactor, MobA, MobB, MoeA and MogA interact with apo-NarG [Vergnes04]. TorD may act as the enzyme-specific chaperone for molybdenum cofactor insertion into TorA [Genest08].

A mobA mutant contains elevated levels of molybdopterin, but no detectable molybdopterin guanine dinucleotide (MGD) [Johnson91].

The mob locus is expressed at a low level under both aerobic and anaerobic growth conditions [IobbiNivol95].

mobA was initially identified as one of several genes that, when mutated, result in resistance to chlorate. Chlorate resistance of the chlB mutants is the result of inactive nitrate reductase, which is a molybdoenzyme [MacGregor72, Amy81].

ChlB: "chlorate resistance"

MobA: "molybdenum cofactor biosynthesis, chlB group, gene A [Shanmugam92]

Reviews: [Rajagopalan97, Rizzi02, IobbiNivol13]

Citations: [Casse70, MacGregor71, Venables72, MacGregor73, Lambdren76, Giordano80, Takagi81, Miller83a, Giordano84, Campbell85, Saracino86, Silvestro86, Iuchi87a, Boxer87, Reiss87, Giordano90, Santini92, Rothery95, Rothery98a]

Locations: cytosol

Map Position: [4,039,438 <- 4,040,022] (87.06 centisomes)
Length: 585 bp / 194 aa

Molecular Weight of Polypeptide: 21.643 kD (from nucleotide sequence), 22.0 kD (experimental) [Palmer94 ]

Unification Links: ASAP:ABE-0012593 , CGSC:921 , DIP:DIP-10233N , EchoBASE:EB1776 , EcoGene:EG11829 , EcoliWiki:b3857 , Mint:MINT-1232345 , ModBase:P32173 , OU-Microarray:b3857 , PortEco:mobA , PR:PRO_000023265 , Pride:P32173 , Protein Model Portal:P32173 , RefSeq:NP_418294 , RegulonDB:EG11829 , SMR:P32173 , String:511145.b3857 , UniProt:P32173

Relationship Links: InterPro:IN-FAMILY:IPR013482 , InterPro:IN-FAMILY:IPR025877 , PDB:Structure:1E5K , PDB:Structure:1FR9 , PDB:Structure:1FRW , PDB:Structure:1H4C , PDB:Structure:1H4D , PDB:Structure:1H4E , PDB:Structure:1HJJ , PDB:Structure:1HJL , Pfam:IN-FAMILY:PF12804

Gene-Reaction Schematic: ?

GO Terms:

Biological Process: GO:1902758 - bis(molybdopterin guanine dinucleotide)molybdenum biosynthetic process Inferred from experiment [Johnson91]
GO:0006777 - Mo-molybdopterin cofactor biosynthetic process Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0019720 - Mo-molybdopterin cofactor metabolic process Inferred by computational analysis [GOA06]
Molecular Function: GO:0000287 - magnesium ion binding Inferred from experiment [Lake00]
GO:0005515 - protein binding Inferred from experiment [Magalon02]
GO:0061603 - molybdenum cofactor guanylyltransferase activity Inferred from experiment Inferred by computational analysis [GOA01, Neumann11]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11a]
GO:0003824 - catalytic activity Inferred by computational analysis [GOA01a]
GO:0005525 - GTP binding Inferred by computational analysis [UniProtGOA11a, GOA06]
GO:0016740 - transferase activity Inferred by computational analysis [UniProtGOA11a]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11a]
GO:0070568 - guanylyltransferase activity Inferred by computational analysis [GOA06]
Cellular Component: GO:0005737 - cytoplasm Inferred by computational analysis [UniProtGOA11, UniProtGOA11a, GOA06]
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: metabolism biosynthesis of building blocks cofactors, small molecule carriers molybdenum

Essentiality data for mobA knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 1]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 2]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 3]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 2]

Credits:
Last-Curated ? 07-Oct-2013 by Keseler I , SRI International


Enzymatic reaction of: molybdenum cofactor guanylyltransferase

Synonyms: MGD synthase, molybdopterin guanine dinucleotide synthase, MoCo guanylyltransferase, GTP:molybdopterin guanylyltransferase

EC Number: 2.7.7.77

MoO2-molybdopterin cofactor + GTP + H+ <=> guanylyl molybdenum cofactor + diphosphate

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the Enzyme Commission system.

The reaction is favored in the direction shown.

In Pathways: guanylyl molybdenum cofactor biosynthesis

Summary:
MobA activity was measured as activation of molybdoenzymes in cell extracts of mobA mutants [Santini92, Palmer94, Palmer96, Guse03].

Cofactors or Prosthetic Groups: Mg2+ [Lake00]

Alternative Cofactors for Mg2+: Mn2+

Kinetic Parameters:

Substrate
Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
Citations
GTP
6.5
0.0048, 0.0048
[Neumann11, BRENDA14]


Sequence Features

Feature Class Location Attached Group Citations Comment
Nucleotide-Phosphate-Binding-Region 12 -> 14 GTP
[UniProt14]
UniProt: GTP.
Mutagenesis-Variant 12 -> 14  
[Neumann11, UniProt12a]
Alternate sequence: LAG → TAA; UniProt: 7.5-fold decrease in affinity for GTP and nearly no effect on catalytic activity. Displays a 3-fold decrease in activity with GTP and gains a low activity with CTP as substrate; when associated with 79-LLTS-82.
Mutagenesis-Variant 15  
[Guse03, UniProt12a]
Alternate sequence: G → L; UniProt: Complete loss of catalytic activity. Still capable of binding MPT and MGD and interacting with both MoeA and MobB.
Mutagenesis-Variant 19  
[Guse03, UniProt12a]
Alternate sequence: R → A; UniProt: Slight reduction in catalytic activity.
Mutagenesis-Variant 22  
[Guse03, UniProt12a]
Alternate sequence: G → L; UniProt: Nearly no effect on catalytic activity.
Mutagenesis-Variant 25  
[Guse03, UniProt12a]
Alternate sequence: K → A; UniProt: Marked reduction in catalytic activity. Still capable of interacting with both MoeA and MobB.
Amino-Acid-Sites-That-Bind 25  
[UniProt12a]
UniProt: GTP.
Amino-Acid-Sites-That-Bind 53  
[UniProt12a]
UniProt: GTP.
Amino-Acid-Sites-That-Bind 71  
[UniProt12a]
UniProt: GTP.
Mutagenesis-Variant 78  
[Guse03, UniProt12a]
Alternate sequence: G → L; UniProt: Nearly no effect on catalytic activity.
Mutagenesis-Variant 79 -> 82  
[Guse03, Neumann11, UniProt12a]
Alternate sequence: PLAG → LLTS; UniProt: 11-fold decrease in affinity for GTP and nearly no effect on catalytic activity. Displays a 3-fold decrease in activity with GTP and gains a low activity with CTP as substrate; when associated with 12-TAA-14.
Mutagenesis-Variant 82  
[Guse03, UniProt12a]
Alternate sequence: G → L; UniProt: Slight reduction in catalytic activity.
Mutagenesis-Variant 101  
[Guse03, UniProt12a]
Alternate sequence: D → N; UniProt: Marked reduction in catalytic activity. Still capable of interacting with both MoeA and MobB.
Alternate sequence: D → A; UniProt: Complete loss of catalytic activity.
Metal-Binding-Site 101  
[UniProt12a]
UniProt: Magnesium.
Mutagenesis-Variant 156  
[Guse03, UniProt12a]
Alternate sequence: R → A; UniProt: Nearly no effect on catalytic activity.
Mutagenesis-Variant 180  
[Guse03, UniProt12a]
Alternate sequence: N → D; UniProt: Nearly no effect on catalytic activity.
Mutagenesis-Variant 182  
[Guse03, UniProt12a]
Alternate sequence: N → D; UniProt: Nearly no effect on catalytic activity.


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b3857 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11829; confirmed by SwissProt match.


References

Amy81: Amy NK (1981). "Identification of the molybdenum cofactor in chlorate-resistant mutants of Escherichia coli." J Bacteriol 148(1);274-82. PMID: 7026535

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

Boxer87: Boxer DH, Low DC, Pommier J, Giordano G (1987). "Involvement of a low-molecular-weight substance in in vitro activation of the molybdoenzyme respiratory nitrate reductase from a chlB mutant of Escherichia coli." J Bacteriol 1987;169(10);4678-85. PMID: 3308848

BRENDA14: BRENDA team (2014). "Imported from BRENDA version existing on Aug 2014." http://www.brenda-enzymes.org.

Campbell85: Campbell AM, del Campillo-Campbell A, Villaret DB (1985). "Molybdate reduction by Escherichia coli K-12 and its chl mutants." Proc Natl Acad Sci U S A 82(1);227-31. PMID: 3881754

Casse70: Casse F (1970). "Mapping of the gene chl-B controlling membran bound nitrate reductase and formic hydrogen-lyase activities in Escherichia coli K 12." Biochem Biophys Res Commun 39(3);429-36. PMID: 4912199

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

Genest08: Genest O, Neumann M, Seduk F, Stocklein W, Mejean V, Leimkuhler S, Iobbi-Nivol C (2008). "Dedicated metallochaperone connects apoenzyme and molybdenum cofactor biosynthesis components." J Biol Chem 283(31);21433-40. PMID: 18522945

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

Giordano80: Giordano G, Grillet L, Pommier J, Terriere C, Haddock BA, Azoulay E (1980). "Precursor forms of the subunits of nitrate reductase in chlA and chlB mutants of Escherichia coli K12." Eur J Biochem 105(2);297-306. PMID: 6991254

Giordano84: Giordano G, Violet M, Medani CL, Pommier J (1984). "A common pathway for the activation of several molybdoenzymes in Escherichia coli K12." Biochim Biophys Acta 798(2);216-25. PMID: 6370312

Giordano90: Giordano G, Boxer DH, Pommier J (1990). "Molybdenum cofactor requirement for in vitro activation of apo-molybdoenzymes of Escherichia coli." Mol Microbiol 4(4);645-50. PMID: 2141097

GOA01: GOA, MGI (2001). "Gene Ontology annotation based on Enzyme Commission mapping." Genomics 74;121-128.

GOA01a: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

Guse03: Guse A, Stevenson CE, Kuper J, Buchanan G, Schwarz G, Giordano G, Magalon A, Mendel RR, Lawson DM, Palmer T (2003). "Biochemical and structural analysis of the molybdenum cofactor biosynthesis protein MobA." J Biol Chem 278(28);25302-7. PMID: 12719427

IobbiNivol13: Iobbi-Nivol C, Leimkuhler S (2013). "Molybdenum enzymes, their maturation and molybdenum cofactor biosynthesis in Escherichia coli." Biochim Biophys Acta 1827(8-9);1086-101. PMID: 23201473

IobbiNivol95: Iobbi-Nivol C, Palmer T, Whitty PW, McNairn E, Boxer DH (1995). "The mob locus of Escherichia coli K12 required for molybdenum cofactor biosynthesis is expressed at very low levels." Microbiology 1995;141 ( Pt 7);1663-71. PMID: 7551035

Iuchi87a: Iuchi S, Lin EC (1987). "Molybdenum effector of fumarate reductase repression and nitrate reductase induction in Escherichia coli." J Bacteriol 169(8);3720-5. PMID: 3301812

Johnson91: Johnson JL, Indermaur LW, Rajagopalan KV (1991). "Molybdenum cofactor biosynthesis in Escherichia coli. Requirement of the chlB gene product for the formation of molybdopterin guanine dinucleotide." J Biol Chem 1991;266(19);12140-5. PMID: 1648082

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

Lake00: Lake MW, Temple CA, Rajagopalan KV, Schindelin H (2000). "The crystal structure of the Escherichia coli MobA protein provides insight into molybdopterin guanine dinucleotide biosynthesis." J Biol Chem 2000;275(51);40211-7. PMID: 10978347

Lambdren76: Lambdren PR, Guest JR (1976). "A novel method for isolating chlorate-resistant mutants of Escherichia coli K12 by anaerobic selection on a lactate plus fumarate medium." J Gen Microbiol 93(1);173-6. PMID: 772165

MacGregor71: MacGregor CH, Schnaitman CA (1971). "Alterations in the cytoplasmic membrane proteins of various chlorate-resistant mutants of Escherichia coli." J Bacteriol 108(1);564-70. PMID: 4941570

MacGregor72: MacGregor CH, Schnaitman CA (1972). "Restoration of reduced nicotinamide adenine dinucleotide phosphate-nitrate reductase activity of a Neurospora mutant by extracts of various chlorate-resistant mutants of Escherichia coli." J Bacteriol 112(1);388-91. PMID: 4404057

MacGregor73: MacGregor CH, Schnaitman CA (1973). "Reconstitution of nitrate reductase activity and formation of membrane particles from cytoplasmic extracts of chlorate-resistant mutants of Escherichia coli." J Bacteriol 114(3);1164-76. PMID: 4576401

Magalon02: Magalon A, Frixon C, Pommier J, Giordano G, Blasco F (2002). "In vivo interactions between gene products involved in the final stages of molybdenum cofactor biosynthesis in Escherichia coli." J Biol Chem 277(50);48199-204. PMID: 12372836

Miller83a: Miller JB, Amy NK (1983). "Molybdenum cofactor in chlorate-resistant and nitrate reductase-deficient insertion mutants of Escherichia coli." J Bacteriol 155(2);793-801. PMID: 6307982

Neumann11: Neumann M, Seduk F, Iobbi-Nivol C, Leimkuhler S (2011). "Molybdopterin dinucleotide biosynthesis in Escherichia coli: identification of amino acid residues of molybdopterin dinucleotide transferases that determine specificity for binding of guanine or cytosine nucleotides." J Biol Chem 286(2);1400-8. PMID: 21081498

Palmer94: Palmer T, Vasishta A, Whitty PW, Boxer DH (1994). "Isolation of protein FA, a product of the mob locus required for molybdenum cofactor biosynthesis in Escherichia coli." Eur J Biochem 1994;222(2);687-92. PMID: 8020507

Palmer96: Palmer T, Santini CL, Iobbi-Nivol C, Eaves DJ, Boxer DH, Giordano G (1996). "Involvement of the narJ and mob gene products in distinct steps in the biosynthesis of the molybdoenzyme nitrate reductase in Escherichia coli." Mol Microbiol 1996;20(4);875-84. PMID: 8793883

Rajagopalan97: Rajagopalan KV (1997). "Biosynthesis and processing of the molybdenum cofactors." Biochem Soc Trans 1997;25(3);757-61. PMID: 9388540

Reiss87: Reiss J, Kleinhofs A, Klingmuller W (1987). "Cloning of seven differently complementing DNA fragments with chl functions from Escherichia coli K12." Mol Gen Genet 206(2);352-5. PMID: 3035339

Reschke13: Reschke S, Sigfridsson KG, Kaufmann P, Leidel N, Horn S, Gast K, Schulzke C, Haumann M, Leimkuhler S (2013). "Identification of a bis-molybdopterin intermediate in molybdenum cofactor biosynthesis in Escherichia coli." J Biol Chem 288(41);29736-45. PMID: 24003231

Rizzi02: Rizzi M, Schindelin H (2002). "Structural biology of enzymes involved in NAD and molybdenum cofactor biosynthesis." Curr Opin Struct Biol 12(6);709-20. PMID: 12504674

Rothery95: Rothery RA, Grant JL, Johnson JL, Rajagopalan KV, Weiner JH (1995). "Association of molybdopterin guanine dinucleotide with Escherichia coli dimethyl sulfoxide reductase: effect of tungstate and a mob mutation." J Bacteriol 1995;177(8);2057-63. PMID: 7721698

Rothery98a: Rothery RA, Magalon A, Giordano G, Guigliarelli B, Blasco F, Weiner JH (1998). "The molybdenum cofactor of Escherichia coli nitrate reductase A (NarGHI). Effect of a mobAB mutation and interactions with [Fe-S] clusters." J Biol Chem 1998;273(13);7462-9. PMID: 9516445

Santini92: Santini CL, Iobbi-Nivol C, Romane C, Boxer DH, Giordano G (1992). "Molybdoenzyme biosynthesis in Escherichia coli: in vitro activation of purified nitrate reductase from a chlB mutant." J Bacteriol 1992;174(24);7934-40. PMID: 1459941

Saracino86: Saracino L, Violet M, Boxer DH, Giordano G (1986). "Activation in vitro of respiratory nitrate reductase of Escherichia coli K12 grown in the presence of tungstate. Involvement of molybdenum cofactor." Eur J Biochem 158(3);483-90. PMID: 3525161

Shanmugam92: Shanmugam KT, Stewart V, Gunsalus RP, Boxer DH, Cole JA, Chippaux M, DeMoss JA, Giordano G, Lin EC, Rajagopalan KV (1992). "Proposed nomenclature for the genes involved in molybdenum metabolism in Escherichia coli and Salmonella typhimurium." Mol Microbiol 6(22);3452-4. PMID: 1484496

Silvestro86: Silvestro A, Pommier J, Giordano G (1986). "Molybdenum cofactor: a compound in the in vitro activation of both nitrate reductase and trimethylamine-N-oxide reductase activities in Escherichia coli K12." Biochim Biophys Acta 872(3);243-52. PMID: 3524687

Stevenson00: Stevenson CE, Sargent F, Buchanan G, Palmer T, Lawson DM (2000). "Crystal structure of the molybdenum cofactor biosynthesis protein MobA from Escherichia coli at near-atomic resolution." Structure 8(11);1115-25. PMID: 11080634

Takagi81: Takagi M, Tsuchiya T, Ishimoto M (1981). "Proton translocation coupled to trimethylamine N-oxide reduction in anaerobically grown Escherichia coli." J Bacteriol 148(3);762-8. PMID: 7031034

UniProt12a: UniProt Consortium (2012). "UniProt version 2012-09 released on 2012-09-12 00:00:00." Database.

UniProt14: UniProt Consortium (2014). "UniProt version 2014-01 released on 2014-01-01 00:00:00." Database.

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on the manual assignment of UniProtKB Subcellular Location terms in UniProtKB/Swiss-Prot entries."

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

Venables72: Venables WA (1972). "Genetic studies with nitrate reductase-less mutants of Escherichia coli. I. Fine structure analysis of the narA, narB and narE loci." Mol Gen Genet 114(3);223-31. PMID: 4552503

Vergnes04: Vergnes A, Gouffi-Belhabich K, Blasco F, Giordano G, Magalon A (2004). "Involvement of the molybdenum cofactor biosynthetic machinery in the maturation of the Escherichia coli nitrate reductase A." J Biol Chem 279(40);41398-403. PMID: 15247236

Other References Related to Gene Regulation

Salmon03: Salmon K, Hung SP, Mekjian K, Baldi P, Hatfield GW, Gunsalus RP (2003). "Global gene expression profiling in Escherichia coli K12. The effects of oxygen availability and FNR." J Biol Chem 278(32);29837-55. PMID: 12754220


Report Errors or Provide Feedback
Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
Page generated by SRI International Pathway Tools version 18.5 on Thu Dec 18, 2014, BIOCYC14A.