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Escherichia coli K-12 substr. MG1655 Enzyme: 4-amino-4-deoxychorismate synthase

Component of: para-aminobenzoate synthase multi-enzyme complex (summary available)

Subunit composition of 4-amino-4-deoxychorismate synthase = [PabB][PabA]

Summary:
E. coli K-12 genes pabA and pabB were previously thought to be responsible for 4-aminobenzoate (para-aminobenzoate) synthesis in a single step [Huang67, Huang70]. Later work revised this, showing that these gene products form a PabAB heterodimeric enzyme that catalyzes the formation of the intermediate 4-amino-4-deoxychorismate. A second enzyme, aminodeoxychorismate lyase encoded by pabC, catalyzes the conversion of this intermediate to 4-aminobenzoate [Nichols89, Ye90, Anderson91, Green91, Green92] (see pathway 4-aminobenzoate biosynthesis). Thus, the name p-aminobenzoate synthase for PabAB is now an historical description [Green91].

PabA and PabB form a 1:1 heterodimeric complex that catalyzes the conversion of chorismate and glutamine to the intermediate 4-amino-4-deoxychorismate. PabA functions as a glutaminase, but only when in complex with PabB. PabA alone is inactive and is therefore a conditional glutaminase that is activated by complex formation with PabB. PabB is an aminodeoxychorismate synthase that utilizes chorismate, and the nacent ammonia derived from glutamine hydrolysis by PabA, as substrates. In the absence of PabA, PabB can utilize added ammonia (at a reduced rate) to aminate chorismate and produce 4-amino-4-deoxychorismate [Roux92, Anderson91, Ye90].

A complex between PabA and PabB that was stable to gel filtration chromatography could not be demonstrated [Roux92], although a later report found gel filtration conditions that demonstrated a complex [Rayl96]. At least a transient complex must form to allow glutamine-dependent production of 4-amino-4-deoxychorismate [Parsons02].

The reaction catalyzed by PabB in vitro in the presence of ammonia is reversible in the absence of PabC. The product 4-amino-4-deoxychorismate was identified by NMR spectroscopy [Anderson91]. Titration of PabA with PabB showed a gain of glutaminase activity that reached a plateau at a 1:1 ratio of PabA/PabB, suggesting that the effect of PabB is as a stoichiometric positive allosteric effector on the PabA subunit [Roux92].

4-Amino-4-deoxychorismate synthase is one of several chorismate-utilizing enzymes in E. coli, along with EntC, UbiC, TyrA, PheA, and trpDE. PabB showed 26% amino acid sequence similarity and 40% nucleotide sequence similarity to trpE, suggesting common ancestry [Goncharoff84].

Gene-Reaction Schematic: ?

Credits:
Last-Curated ? 07-May-2012 by Fulcher C , SRI International


Enzymatic reaction of: 4-amino-4-deoxychorismate synthase

Synonyms: para-aminobenzoate synthase, p-aminobenzoate synthase, PABA synthase, PABS, aminodeoxychorismate synthase

EC Number: 2.6.1.85

L-glutamine + chorismate <=> 4-amino-4-deoxychorismate + L-glutamate

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.

This reaction is reversible.

Alternative Substrates [Comment 1]:

In Pathways: superpathway of chorismate metabolism , superpathway of tetrahydrofolate biosynthesis , 4-aminobenzoate biosynthesis

Cofactors or Prosthetic Groups: Mg2+ [Comment 2, Ye90]

Inhibitors (Unknown Mechanism): 6-diazo-5-oxonorleucine [Roux92, Comment 3] , cycloheptadienyl [Walsh87, Comment 4]

Kinetic Parameters:

Substrate
Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
Citations
chorismate
4.2
[Viswanathan95]
chorismate
13.0, 58.0
0.032, 0.53
[He04, BRENDA14]
L-glutamine
1600.0
[Viswanathan95]
L-glutamine
0.67
[Ziebart10, BRENDA14]


Subunit of: para-aminobenzoate synthase multi-enzyme complex

Subunit composition of para-aminobenzoate synthase multi-enzyme complex = [(PabB)(PabA)][(PabC)2]
         4-amino-4-deoxychorismate synthase = (PabB)(PabA) (extended summary available)
         aminodeoxychorismate lyase = (PabC)2 (summary available)

Summary:
The three proteins PabA, PabB and PabC have been functionally described as three subunits of a 4-aminobenzoate (p-aminobenzoate) synthase multienzyme complex [Roux92]. Although no such complex has been isolated, the possibility of a ternary complex in vivo remains [Green92]. Together these proteins catalyze the conversion of glutamine and chorismate to 4-aminobenzoate, glutamate and pyruvate (see pathway 4-aminobenzoate biosynthesis).

Credits:
Last-Curated ? 07-May-2012 by Fulcher C , SRI International


Subunit of 4-amino-4-deoxychorismate synthase: PabB

Synonyms: para-aminobenzoate synthase component I, ADC synthase

Gene: pabB Accession Numbers: EG10683 (EcoCyc), b1812, ECK1810

Locations: cytosol

Sequence Length: 453 AAs

Molecular Weight: 50.97 kD (from nucleotide sequence)

pI: 5.32

GO Terms:

Biological Process: GO:0046654 - tetrahydrofolate biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA12, He06]
GO:0009058 - biosynthetic process Inferred by computational analysis [GOA01a]
GO:0009396 - folic acid-containing compound biosynthetic process Inferred by computational analysis [GOA01a]
GO:0046656 - folic acid biosynthetic process Inferred by computational analysis [UniProtGOA11a]
Molecular Function: GO:0000287 - magnesium ion binding Inferred from experiment [Ye90]
GO:0046820 - 4-amino-4-deoxychorismate synthase activity Inferred from experiment Inferred by computational analysis [GOA01, Ye90, Viswanathan95]
GO:0016740 - transferase activity Inferred by computational analysis [UniProtGOA11a]
GO:0016833 - oxo-acid-lyase activity Inferred by computational analysis [GOA01a]
Cellular Component: GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

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

Unification Links: DIP:DIP-10434N , EcoliWiki:b1812 , ModBase:P05041 , PR:PRO_000023485 , Pride:P05041 , Protein Model Portal:P05041 , RefSeq:NP_416326 , SMR:P05041 , String:511145.b1812 , UniProt:P05041

Relationship Links: InterPro:IN-FAMILY:IPR005801 , InterPro:IN-FAMILY:IPR005802 , InterPro:IN-FAMILY:IPR006805 , InterPro:IN-FAMILY:IPR015890 , InterPro:IN-FAMILY:IPR019999 , PDB:Structure:1K0E , PDB:Structure:1K0G , Pfam:IN-FAMILY:PF00425 , Pfam:IN-FAMILY:PF04715 , Prints:IN-FAMILY:PR00095

Summary:
Component I catalyzes the formation of 4-amino-4-deoxychorismate (ADC) by binding chorismate and ammonia.

The crystal structure of PabB has been determined at 2.0 Å resolution [Parsons02].

Essentiality data for pabB knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 5]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 6]
M9 medium with 0.4% glucose No 37 Aerobic 7.2 0.27 No [Patrick07, Comment 7]
M9 medium with 1% glycerol No 37 Aerobic 7.2 0.35 No [Joyce06]
MOPS medium with 0.4% glucose Indeterminate 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 6]
No [Feist07, Comment 8]

Subunit of 4-amino-4-deoxychorismate synthase: PabA

Synonyms: para-aminobenzoate synthase glutamine amidotransferase component II

Gene: pabA Accession Numbers: EG10682 (EcoCyc), b3360, ECK3348

Locations: cytosol

Sequence Length: 187 AAs

Molecular Weight: 20.772 kD (from nucleotide sequence)

pI: 6.52

GO Terms:

Biological Process: GO:0046654 - tetrahydrofolate biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA12, Huang70]
GO:0006541 - glutamine metabolic process Inferred by computational analysis [UniProtGOA11a]
GO:0008152 - metabolic process Inferred by computational analysis [GOA01a]
GO:0046656 - folic acid biosynthetic process Inferred by computational analysis [UniProtGOA11a]
Molecular Function: GO:0046820 - 4-amino-4-deoxychorismate synthase activity Inferred from experiment Inferred by computational analysis [GOA01, Huang70]
GO:0016740 - transferase activity Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

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

Unification Links: DIP:DIP-10433N , EcoliWiki:b3360 , ModBase:P00903 , PR:PRO_000023484 , Protein Model Portal:P00903 , RefSeq:NP_417819 , SMR:P00903 , String:511145.b3360 , Swiss-Model:P00903 , UniProt:P00903

Relationship Links: InterPro:IN-FAMILY:IPR006221 , InterPro:IN-FAMILY:IPR017926 , Pfam:IN-FAMILY:PF00117 , Prosite:IN-FAMILY:PS51273

Summary:
Component II provides the glutamine amidotransferase activity.

Analysis of PabA by site-directed mutagenesis suggested a role for the conserved Cys-79 as a catalytically active residue [Roux93].

Essentiality data for pabA knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 5]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 6]
M9 medium with 0.4% glucose No 37 Aerobic 7.2 0.27 No [Patrick07, Comment 7]
M9 medium with 1% glycerol No 37 Aerobic 7.2 0.35 No [Joyce06]
MOPS medium with 0.4% glucose Indeterminate 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 6]
No [Feist07, Comment 8]

References

Anderson91: Anderson KS, Kati WM, Ye Q-Z, Liu J, Walsh CT, Benesi AJ (1991). "Isolation and structure elucidation of the 4-amino-4-deoxychorismate intermediate in the PABA enzymatic pathway." J. Am. Chem. Soc. 113, 3198-3200.

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

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

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

Feist07: Feist AM, Henry CS, Reed JL, Krummenacker M, Joyce AR, Karp PD, Broadbelt LJ, Hatzimanikatis V, Palsson BO (2007). "A genome-scale metabolic reconstruction for Escherichia coli K-12 MG1655 that accounts for 1260 ORFs and thermodynamic information." Mol Syst Biol 3;121. PMID: 17593909

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

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."

Goncharoff84: Goncharoff P, Nichols BP (1984). "Nucleotide sequence of Escherichia coli pabB indicates a common evolutionary origin of p-aminobenzoate synthetase and anthranilate synthetase." J Bacteriol 1984;159(1);57-62. PMID: 6330050

Green91: Green JM, Nichols BP (1991). "p-Aminobenzoate biosynthesis in Escherichia coli. Purification of aminodeoxychorismate lyase and cloning of pabC." J Biol Chem 1991;266(20);12971-5. PMID: 2071583

Green92: Green JM, Merkel WK, Nichols BP (1992). "Characterization and sequence of Escherichia coli pabC, the gene encoding aminodeoxychorismate lyase, a pyridoxal phosphate-containing enzyme." J Bacteriol 1992;174(16);5317-23. PMID: 1644759

He04: He Z, Stigers Lavoie KD, Bartlett PA, Toney MD (2004). "Conservation of mechanism in three chorismate-utilizing enzymes." J Am Chem Soc 126(8);2378-85. PMID: 14982443

He06: He Z, Toney MD (2006). "Direct detection and kinetic analysis of covalent intermediate formation in the 4-amino-4-deoxychorismate synthase catalyzed reaction." Biochemistry 45(15);5019-28. PMID: 16605270

Huang67: Huang M, Pittard J (1967). "Genetic analysis of mutant strains of Escherichia coli requiring p-aminobenzoic acid for growth." J Bacteriol 93(6);1938-42. PMID: 5337773

Huang70: Huang M, Gibson F (1970). "Biosynthesis of 4-aminobenzoate in Escherichia coli." J Bacteriol 102(3);767-73. PMID: 4914080

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

Nichols89: Nichols BP, Seibold AM, Doktor SZ (1989). "para-aminobenzoate synthesis from chorismate occurs in two steps." J Biol Chem 264(15);8597-601. PMID: 2656685

Parsons02: Parsons JF, Jensen PY, Pachikara AS, Howard AJ, Eisenstein E, Ladner JE (2002). "Structure of Escherichia coli aminodeoxychorismate synthase: architectural conservation and diversity in chorismate-utilizing enzymes." Biochemistry 41(7);2198-208. PMID: 11841211

Patrick07: Patrick WM, Quandt EM, Swartzlander DB, Matsumura I (2007). "Multicopy suppression underpins metabolic evolvability." Mol Biol Evol 24(12);2716-22. PMID: 17884825

Rayl96: Rayl EA, Green JM, Nichols BP (1996). "Escherichia coli aminodeoxychorismate synthase: analysis of pabB mutations affecting catalysis and subunit association." Biochim Biophys Acta 1295(1);81-8. PMID: 8679677

Roux92: Roux B, Walsh CT (1992). "p-aminobenzoate synthesis in Escherichia coli: kinetic and mechanistic characterization of the amidotransferase PabA." Biochemistry 1992;31(30);6904-10. PMID: 1637823

Roux93: Roux B, Walsh CT (1993). "p-Aminobenzoate synthesis in Escherichia coli: mutational analysis of three conserved amino acid residues of the amidotransferase PabA." Biochemistry 1993;32(14);3763-8. PMID: 8096767

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

UniProtGOA12: UniProt-GOA (2012). "Gene Ontology annotation based on UniPathway vocabulary mapping."

Viswanathan95: Viswanathan VK, Green JM, Nichols BP (1995). "Kinetic characterization of 4-amino 4-deoxychorismate synthase from Escherichia coli." J Bacteriol 177(20);5918-23. PMID: 7592344

Walsh87: Walsh CT, Erion MD, Walts AE, Delany JJ, Berchtold GA (1987). "Chorismate aminations: partial purification of Escherichia coli PABA synthase and mechanistic comparison with anthranilate synthase." Biochemistry 1987;26(15);4734-45. PMID: 3311153

Ye90: Ye QZ, Liu J, Walsh CT (1990). "p-Aminobenzoate synthesis in Escherichia coli: purification and characterization of PabB as aminodeoxychorismate synthase and enzyme X as aminodeoxychorismate lyase." Proc Natl Acad Sci U S A 1990;87(23);9391-5. PMID: 2251281

Ziebart10: Ziebart KT, Toney MD (2010). "Nucleophile specificity in anthranilate synthase, aminodeoxychorismate synthase, isochorismate synthase, and salicylate synthase." Biochemistry 49(13);2851-9. PMID: 20170126


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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 Mon Dec 22, 2014, biocyc11.