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Escherichia coli K-12 substr. MG1655 Enzyme: porphobilinogen synthase



Gene: hemB Accession Numbers: EG10428 (EcoCyc), b0369, ECK0366

Synonyms: ncf

Regulation Summary Diagram: ?

Subunit composition of porphobilinogen synthase = [HemB]8
         porphobilinogen synthase = HemB

Summary:
Porphobilinogen synthase (PBGS) catalyzes the synthesis of porphobilinogen from two molecules of δ-aminolevulinate via a Schiff-base intermediate. PBGS is a metalloenzyme that requires Zn2+ for activity [Spencer93]. Mg2+ is not required for activity, however, addition may result in a 2-4 fold stimulation of the Zn-induced PBGS [Spencer93, Mitchell93, Jaffe95].

The crystal structures of PBGS and PBGS complexed with an inhibitor have been resolved at 2.0 and 1.9 Å, respectively [Erskine99, Kervinen01, Jaffe02]. Four dimers interact to form an octamer in which each active site is located on the surface. The active site contains two lysine residues, one of which forms a Schiff-base link with the bound substrate analogue, 5-aminolevulinic acid. Mutagenesis studies of this conserved lysine showed that PBGS was disabled, but not inactivated [Mitchell01].

There are two substrate binding sites, the P-site and A-site. The P-site binds the 5-aminolevulinic acid, and the A-site recognizes the 5-aminolevulinic acid molecule that forms the side of porphobilinogen carrying the acetic acid side chain [Erskine99, ShoolinginJorda02]. Inhibitors, including those which act as competitive inhibitors by forming a Schiff base at the P-site, have been extensively studied [ShoolinginJorda02, Jarret00].

Mutants of hemB had no 5-aminolevulinate dehydratase and extremely low porphobilinogen deaminase activity [Umanoff88].

Locations: cytosol

Map Position: [387,977 <- 388,951] (8.36 centisomes)
Length: 975 bp / 324 aa

Molecular Weight of Polypeptide: 35.625 kD (from nucleotide sequence), 36.5 kD (experimental) [Spencer93 ]

Molecular Weight of Multimer: 270.0 kD (experimental) [Spencer93]

pI: 5.39, 5.0 [Spencer93]

Unification Links: ASAP:ABE-0001266 , CGSC:647 , DIP:DIP-36209N , EchoBASE:EB0423 , EcoGene:EG10428 , EcoliWiki:b0369 , Mint:MINT-6765615 , ModBase:P0ACB2 , OU-Microarray:b0369 , PortEco:hemB , Pride:P0ACB2 , Protein Model Portal:P0ACB2 , RefSeq:NP_414903 , RegulonDB:EG10428 , SMR:P0ACB2 , String:511145.b0369 , UniProt:P0ACB2

Relationship Links: InterPro:IN-FAMILY:IPR001731 , InterPro:IN-FAMILY:IPR013785 , Panther:IN-FAMILY:PTHR11458 , PDB:Structure:1B4E , PDB:Structure:1I8J , PDB:Structure:1L6S , PDB:Structure:1L6Y , Pfam:IN-FAMILY:PF00490 , Prints:IN-FAMILY:PR00144 , Prosite:IN-FAMILY:PS00169 , Smart:IN-FAMILY:SM01004

Gene-Reaction Schematic: ?

Genetic Regulation Schematic: ?

GO Terms:

Biological Process: GO:0006782 - protoporphyrinogen IX biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA12, Nishimura93a]
GO:0006783 - heme biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA11a, Li88a]
GO:0006779 - porphyrin-containing compound biosynthetic process Inferred by computational analysis [UniProtGOA11a]
GO:0008152 - metabolic process Inferred by computational analysis [UniProtGOA11a]
GO:0033014 - tetrapyrrole biosynthetic process Inferred by computational analysis [GOA01a]
Molecular Function: GO:0004655 - porphobilinogen synthase activity Inferred from experiment Inferred by computational analysis [GOA01, GOA01a, Spencer93, Li88a]
GO:0008270 - zinc ion binding Inferred from experiment [Spencer93]
GO:0042802 - identical protein binding Inferred from experiment [Spencer93]
GO:0046872 - metal ion binding Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA01a, Spencer93]
GO:0003824 - catalytic activity Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0016829 - lyase activity Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, Lasserre06]

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

Essentiality data for hemB knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB Lennox No 37 Aerobic 7   No [Baba06, Comment 1]

Credits:
Last-Curated ? 10-Jul-2013 by Kubo A , SRI International


Enzymatic reaction of: porphobilinogen synthase

Synonyms: δ-aminolevulinic acid dehydratase, aminolevulinate dehydratase, ALAD, ALADH, PBGS, 5-aminolevulinate hydro-lyase (adding 5-aminolevulinate and cyclizing), 5-aminolevulinate dehydratase

EC Number: 4.2.1.24

2 5-aminolevulinate <=> porphobilinogen + H+ + 2 H2O

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: tetrapyrrole biosynthesis I (from glutamate)

Cofactors or Prosthetic Groups: Zn2+ [Comment 2, Spencer93]

Regulators of Unknown Type: o-phenanthroline [Mitchell93] , Mg2+ [Spencer93]

Kinetic Parameters:

Substrate
Km (μM)
Citations
5-aminolevulinate
1000.0, 800.0
[Spencer93, BRENDA14]

pH(opt) (forward direction): 8.5 [Spencer93]

pH(opt): 7.5 [BRENDA14, Mitchell93]


Sequence Features

Feature Class Location Citations Comment
Cleavage-of-Initial-Methionine 1
[Spencer93, UniProt11]
UniProt: Removed.
Chain 2 -> 324
[UniProt09]
UniProt: Delta-aminolevulinic acid dehydratase;
Sequence-Conflict 18 -> 19
[Li89a, UniProt10a]
Alternate sequence: RA → PR; UniProt: (in Ref. 1; AAA62629);
Sequence-Conflict 19 -> 42
[Echelard88, UniProt10a]
Alternate sequence: AMFEETTLSLNDLVLPIFVEEEID → VCLKRQHLSLTTWCCRSLLKKKLT; UniProt: (in Ref. 2; CAA35467);
Sequence-Conflict 90 -> 91
[Li89a, UniProt10a]
Alternate sequence: SD → ER; UniProt: (in Ref. 1; AAA62629);
Sequence-Conflict 104
[Li89a, UniProt10a]
Alternate sequence: R → P; UniProt: (in Ref. 1; AAA62629);
Metal-Binding-Site 120
[UniProt10a]
UniProt: Zinc; catalytic;
Metal-Binding-Site 122
[UniProt10a]
UniProt: Zinc; catalytic;
Mutagenesis-Variant 127
[Mitchell95, UniProt11]
Alternate sequence: H → A; UniProt: No significant effect on activity; when associated with A-129.
Mutagenesis-Variant 129
[Mitchell95, UniProt11]
Alternate sequence: H → A; UniProt: No significant effect on activity; when associated with A-127.
Metal-Binding-Site 130
[UniProt10a]
UniProt: Zinc; catalytic;
Active-Site 195
[Jaffe02, Kervinen01, UniProt11]
UniProt: Schiff-base intermediate with substrate.
Amino-Acid-Sites-That-Bind 205
[UniProt10a]
UniProt: Substrate 1;
Amino-Acid-Sites-That-Bind 216
[UniProt10a]
UniProt: Substrate 1;
Sequence-Conflict 226
[Echelard88, UniProt10a]
Alternate sequence: R → P; UniProt: (in Ref. 2; CAA35467);
Sequence-Conflict 227
[Li89a, UniProt10a]
Alternate sequence: R → A; UniProt: (in Ref. 1; AAA62629);
Sequence-Conflict 229
[Li89a, UniProt10a]
Alternate sequence: A → G; UniProt: (in Ref. 1; AAA62629);
Sequence-Conflict 231
[Li89a, UniProt10a]
Alternate sequence: R → A; UniProt: (in Ref. 1; AAA62629);
Metal-Binding-Site 232
[UniProt10a]
UniProt: Magnesium;
Sequence-Conflict 233
[Li89a, UniProt10a]
Alternate sequence: S → Y; UniProt: (in Ref. 1; AAA62629);
Sequence-Conflict 241
[Li89a, UniProt10a]
Alternate sequence: A → P; UniProt: (in Ref. 1; AAA62629);
Active-Site 247
[Jaffe02, Kervinen01, UniProt11]
UniProt: Schiff-base intermediate with substrate.
Sequence-Conflict 254
[Li89a, UniProt10a]
Alternate sequence: D → N; UniProt: (in Ref. 1; AAA62629);
Amino-Acid-Sites-That-Bind 273
[UniProt10a]
UniProt: Substrate 2;
Amino-Acid-Sites-That-Bind 312
[UniProt10a]
UniProt: Substrate 2;


Gene Local Context (not to scale): ?

Transcription Unit:

Notes:

History:
Peter D. Karp on Wed Jan 18, 2006:
Gene right-end position adjusted based on analysis performed in the 2005 E. coli annotation update [Riley06 ].
10/20/97 Gene b0369 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10428; confirmed by SwissProt match.


References

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

Echelard88: Echelard Y, Dymetryszyn J, Drolet M, Sasarman A (1988). "Nucleotide sequence of the hemB gene of Escherichia coli K12." Mol Gen Genet 214(3);503-8. PMID: 2464127

Erskine99: Erskine PT, Norton E, Cooper JB, Lambert R, Coker A, Lewis G, Spencer P, Sarwar M, Wood SP, Warren MJ, Shoolingin-Jordan PM (1999). "X-ray structure of 5-aminolevulinic acid dehydratase from Escherichia coli complexed with the inhibitor levulinic acid at 2.0 A resolution." Biochemistry 38(14);4266-76. PMID: 10194344

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."

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

Jaffe02: Jaffe EK, Kervinen J, Martins J, Stauffer F, Neier R, Wlodawer A, Zdanov A (2002). "Species-specific inhibition of porphobilinogen synthase by 4-oxosebacic acid." J Biol Chem 277(22);19792-9. PMID: 11909869

Jaffe95: Jaffe EK, Ali S, Mitchell LW, Taylor KM, Volin M, Markham GD (1995). "Characterization of the role of the stimulatory magnesium of Escherichia coli porphobilinogen synthase." Biochemistry 1995;34(1);244-51. PMID: 7819203

Jarret00: Jarret C, Stauffer F, Henz ME, Marty M, Luond RM, Bobalova J, Schurmann P, Neier R (2000). "Inhibition of Escherichia coli porphobilinogen synthase using analogs of postulated intermediates." Chem Biol 7(3);185-96. PMID: 10712932

Kervinen01: Kervinen J, Jaffe EK, Stauffer F, Neier R, Wlodawer A, Zdanov A (2001). "Mechanistic basis for suicide inactivation of porphobilinogen synthase by 4,7-dioxosebacic acid, an inhibitor that shows dramatic species selectivity." Biochemistry 40(28);8227-36. PMID: 11444968

Lasserre06: Lasserre JP, Beyne E, Pyndiah S, Lapaillerie D, Claverol S, Bonneu M (2006). "A complexomic study of Escherichia coli using two-dimensional blue native/SDS polyacrylamide gel electrophoresis." Electrophoresis 27(16);3306-21. PMID: 16858726

Li88a: Li JM, Umanoff H, Proenca R, Russell CS, Cosloy SD (1988). "Cloning of the Escherichia coli K-12 hemB gene." J Bacteriol 170(2);1021-5. PMID: 3276659

Li89a: Li JM, Russell CS, Cosloy SD (1989). "The structure of the Escherichia coli hemB gene." Gene 75(1);177-84. PMID: 2656410

Mitchell01: Mitchell LW, Volin M, Martins J, Jaffe EK (2001). "Mechanistic implications of mutations to the active site lysine of porphobilinogen synthase." J Biol Chem 276(2);1538-44. PMID: 11032841

Mitchell93: Mitchell LW, Jaffe EK (1993). "Porphobilinogen synthase from Escherichia coli is a Zn(II) metalloenzyme stimulated by Mg(II)." Arch Biochem Biophys 1993;300(1);169-77. PMID: 8424649

Mitchell95: Mitchell LW, Volin M, Jaffe EK (1995). "The phylogenetically conserved histidines of Escherichia coli porphobilinogen synthase are not required for catalysis." J Biol Chem 270(41);24054-9. PMID: 7592604

Nishimura93a: Nishimura K, Nakayashiki T, Inokuchi H (1993). "Cloning and sequencing of the hemE gene encoding uroporphyrinogen III decarboxylase (UPD) from Escherichia coli K-12." Gene 133(1);109-13. PMID: 8224882

Riley06: Riley M, Abe T, Arnaud MB, Berlyn MK, Blattner FR, Chaudhuri RR, Glasner JD, Horiuchi T, Keseler IM, Kosuge T, Mori H, Perna NT, Plunkett G, Rudd KE, Serres MH, Thomas GH, Thomson NR, Wishart D, Wanner BL (2006). "Escherichia coli K-12: a cooperatively developed annotation snapshot--2005." Nucleic Acids Res 34(1);1-9. PMID: 16397293

ShoolinginJorda02: Shoolingin-Jordan PM, Spencer P, Sarwar M, Erskine PE, Cheung KM, Cooper JB, Norton EB (2002). "5-Aminolaevulinic acid dehydratase: metals, mutants and mechanism." Biochem Soc Trans 30(4);584-90. PMID: 12196142

Spencer93: Spencer P, Jordan PM (1993). "Purification and characterization of 5-aminolaevulinic acid dehydratase from Escherichia coli and a study of the reactive thiols at the metal-binding domain." Biochem J 1993;290 ( Pt 1);279-87. PMID: 8439296

Umanoff88: Umanoff H, Russell CS, Cosloy SD (1988). "Availability of porphobilinogen controls appearance of porphobilinogen deaminase activity in Escherichia coli K-12." J Bacteriol 170(10);4969-71. PMID: 3049558

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt10a: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

UniProt11: UniProt Consortium (2011). "UniProt version 2011-06 released on 2011-06-30 00:00:00." Database.

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."

Other References Related to Gene Regulation

Li13: Li F, Wang Y, Gong K, Wang Q, Liang Q, Qi Q (2013). "Constitutive expression of RyhB regulates the heme biosynthesis pathway and increases the 5-aminolevulinic acid accumulation in Escherichia coli." FEMS Microbiol Lett. PMID: 24188714


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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 Sun Dec 21, 2014, biocyc11.