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Escherichia coli K-12 substr. MG1655 Enzyme: cyanase



Gene: cynS Accession Numbers: EG10175 (EcoCyc), b0340, ECK0337

Synonyms: cnt, cyaN

Regulation Summary Diagram: ?

Subunit composition of cyanase = [(CynS)2]5
         cyanase dimer = (CynS)2
                 cyanase monomer = CynS

Summary:
Cyanase is an inducible enzyme in E. coli and was first reported and subsequently studied in E. coli B [Taussig60, Anderson80]. The cyanase operon may have evolved to function in detoxification/decomposition of cyanate arising from intra- and extracellular activities [Anderson90].

It was shown that bicarbonate (HCO3-) is a substrate of the enzyme, and that water does not participate as a substrate [Johnson87a]. The initial product carbamate is unstable and spontaneously decomposes to ammonia and carbon dioxide [Anderson80]. The enzyme shows competitive substrate inhibition by both substrates [Anderson86].

The native enzyme from E. coli B has a molecular weight of ca. 150 kDa and thus appears to be a homodecamer in solution [Anderson80]. A crystal structure of cyanase was solved at 1.65 Å resolution; it showed a homodecamer composed of five dimers. The structure allowed identification of the active site, which is located between dimers and is comprised of residues from four adjacent subunits [Walsh00].

The free sulfhydryl group of the single cysteine residue is not required for catalytic activity, but plays a role in stabilizing the decameric structure [Anderson88c, Anderson94]. The single histidine residue is also required for the stability of the decamer and contributes to its catalytic properties [Anderson94, Anderson94a].

A mutant lacking cyanase is more sensitive to cyanate than the wild type [Guilloton87]; a cynS mutant is not able to use cyanate as the sole source of nitrogen [Sung87].

Gene Citations: [Sung88a, Lamblin94]

Locations: cytosol

Map Position: [358,713 -> 359,183] (7.73 centisomes)
Length: 471 bp / 156 aa

Molecular Weight of Polypeptide: 17.049 kD (from nucleotide sequence), 16.5 kD (experimental) [Sung87 ]

Molecular Weight of Multimer cyanase: 150 kD (experimental) [Anderson80]

pI: 5.29

Unification Links: ASAP:ABE-0001173 , CGSC:15267 , DIP:DIP-9365N , EchoBASE:EB0172 , EcoGene:EG10175 , EcoliWiki:b0340 , ModBase:P00816 , OU-Microarray:b0340 , PortEco:cynS , PR:PRO_000022366 , Pride:P00816 , Protein Model Portal:P00816 , RefSeq:NP_414874 , RegulonDB:EG10175 , SMR:P00816 , String:511145.b0340 , UniProt:P00816

Relationship Links: InterPro:IN-FAMILY:IPR003712 , InterPro:IN-FAMILY:IPR008076 , InterPro:IN-FAMILY:IPR010982 , PDB:Structure:1DW9 , PDB:Structure:1DWK , PDB:Structure:2IU7 , PDB:Structure:2IUO , PDB:Structure:2IV1 , PDB:Structure:2IVB , PDB:Structure:2IVG , PDB:Structure:2IVQ , Pfam:IN-FAMILY:PF02560 , Prints:IN-FAMILY:PR01693

Gene-Reaction Schematic: ?

Genetic Regulation Schematic: ?

GO Terms:

Biological Process: GO:0009440 - cyanate catabolic process Inferred from experiment [Guilloton87]
GO:0009439 - cyanate metabolic process Inferred by computational analysis [GOA06, GOA01a]
Molecular Function: GO:0008824 - cyanate hydratase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01, GOA01a, Anderson87a]
GO:0003677 - DNA binding Inferred by computational analysis [GOA01a]
GO:0016829 - lyase activity Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005737 - cytoplasm Inferred from experiment [Kozliak94]
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: cell processes protection detoxification
metabolism central intermediary metabolism cyanate catabolism
metabolism metabolism of other compounds nitrogen metabolism

Essentiality data for cynS knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 1]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 2]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 1]
Yes [Feist07, Comment 3]

Credits:
Created 27-Mar-2007 by Keseler I , SRI International
Last-Curated ? 27-Mar-2007 by Keseler I , SRI International


Enzymatic reaction of: cyanase

Synonyms: cyanate hydrolase, cyanate C-N-lyase, cyanate aminohydrolase, cyanate lyase, cyanate hydratase

EC Number: 4.2.1.104

cyanate + hydrogen carbonate + H+ <=> carbamate + CO2

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

The reaction is physiologically favored in the direction shown.

In Pathways: cyanate degradation

Summary:
The enzyme was initially purified from E. coli B [Anderson80, Chin83, Anderson86]. A number of kinetic and physical properties of the K-12 and B enzymes were compared and found to be identical [Sung87a].

Cyanase is inhibited by dianions such as oxalate and malonate [Anderson87a].

Inhibitors (Competitive): oxalate [Anderson87a, Comment 4] , formate [Little87, Comment 5] , nitrite [Little87, Comment 5] , nitrate [Little87, Comment 6] , bromide [Little87, Comment 5] , chloride [Little87, Comment 5] , acetate [Little87, Comment 7] , azide [Little87, Comment 8] , cyanate [Anderson86] , hydrogen carbonate [Anderson86]

Kinetic Parameters:

Substrate
Km (μM)
Citations
hydrogen carbonate
950.0
[Anderson94]
cyanate
400.0
[Anderson94]


Sequence Features

Feature Class Location Citations Comment
Sequence-Conflict 34
[Sung88a, UniProt10]
Alternate sequence: D → N; UniProt: (in Ref. 3);
Sequence-Conflict 68
[Sung88a, UniProt10]
Alternate sequence: D → N; UniProt: (in Ref. 3);
Sequence-Conflict 74
[Sung88a, UniProt10]
Alternate sequence: L → S; UniProt: (in Ref. 3);
Active-Site 96
[UniProt10]
Active-Site 99
[UniProt10]
Active-Site 122
[UniProt10]
Sequence-Conflict 125
[Sung88a, UniProt10]
Alternate sequence: N → D; UniProt: (in Ref. 3);


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b0340 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10175; confirmed by SwissProt match.


References

Anderson80: Anderson PM (1980). "Purification and properties of the inducible enzyme cyanase." Biochemistry 19(13);2882-8. PMID: 6994799

Anderson86: Anderson PM, Little RM (1986). "Kinetic properties of cyanase." Biochemistry 25(7);1621-6. PMID: 3518792

Anderson87a: Anderson PM, Johnson WV, Endrizzi JA, Little RM, Korte JJ (1987). "Interaction of mono- and dianions with cyanase: evidence for apparent half-site binding." Biochemistry 26(13);3938-43. PMID: 3651424

Anderson88c: Anderson PM, Johnson WV, Korte JJ, Xiong XF, Sung YC, Fuchs JA (1988). "Reversible dissociation of active octamer of cyanase to inactive dimer promoted by alteration of the sulfhydryl group." J Biol Chem 263(12);5674-80. PMID: 3128546

Anderson90: Anderson PM, Sung YC, Fuchs JA (1990). "The cyanase operon and cyanate metabolism." FEMS Microbiol Rev 1990;7(3-4);247-52. PMID: 2094285

Anderson94: Anderson PM, Korte JJ, Holcomb TA, Cho YG, Son CM, Sung YC (1994). "Formation of intersubunit disulfide bonds and properties of the single histidine and cysteine residues in each subunit relative to the decameric structure of cyanase." J Biol Chem 269(21);15036-45. PMID: 8195141

Anderson94a: Anderson PM, Korte JJ, Holcomb TA (1994). "Reaction of the N-terminal methionine residues in cyanase with diethylpyrocarbonate." Biochemistry 1994;33(47);14121-5. PMID: 7947823

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

Chin83: Chin CC, Anderson PM, Wold F (1983). "The amino acid sequence of Escherichia coli cyanase." J Biol Chem 258(1);276-82. PMID: 6336748

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

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."

Guilloton87: Guilloton M, Karst F (1987). "Isolation and characterization of Escherichia coli mutants lacking inducible cyanase." J Gen Microbiol 133(3);645-53. PMID: 3309165

Johnson87a: Johnson WV, Anderson PM (1987). "Bicarbonate is a recycling substrate for cyanase." J Biol Chem 262(19);9021-5. PMID: 3110153

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

Kozliak94: Kozliak EI, Guilloton MB, Gerami-Nejad M, Fuchs JA, Anderson PM (1994). "Expression of proteins encoded by the Escherichia coli cyn operon: carbon dioxide-enhanced degradation of carbonic anhydrase." J Bacteriol 176(18);5711-7. PMID: 8083164

Lamblin94: Lamblin AF, Fuchs JA (1994). "Functional analysis of the Escherichia coli K-12 cyn operon transcriptional regulation." J Bacteriol 176(21);6613-22. PMID: 7961413

Little87: Little RM, Anderson PM (1987). "Structural properties of cyanase. Denaturation, renaturation, and role of sulfhydryls and oligomeric structure in catalytic activity." J Biol Chem 1987;262(21);10120-6. PMID: 3301828

Sung87: Sung YC, Parsell D, Anderson PM, Fuchs JA (1987). "Identification, mapping, and cloning of the gene encoding cyanase in Escherichia coli K-12." J Bacteriol 169(6);2639-42. PMID: 3034861

Sung87a: Sung YC, Anderson PM, Fuchs JA (1987). "Characterization of high-level expression and sequencing of the Escherichia coli K-12 cynS gene encoding cyanase." J Bacteriol 169(11);5224-30. PMID: 2822670

Sung88a: Sung YC, Fuchs JA (1988). "Characterization of the cyn operon in Escherichia coli K12." J Biol Chem 263(29);14769-75. PMID: 3049588

Taussig60: Taussig A (1960). "The synthesis of the induced enzyme, ''cyanase'', in E. coli." Biochim Biophys Acta 44;510-9. PMID: 13775509

UniProt10: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

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

Walsh00: Walsh MA, Otwinowski Z, Perrakis A, Anderson PM, Joachimiak A (2000). "Structure of cyanase reveals that a novel dimeric and decameric arrangement of subunits is required for formation of the enzyme active site." Structure 8(5);505-14. PMID: 10801492

Other References Related to Gene Regulation

Lamblin93: Lamblin AF, Fuchs JA (1993). "Expression and purification of the cynR regulatory gene product: CynR is a DNA-binding protein." J Bacteriol 1993;175(24);7990-9. PMID: 8253686


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 Fri Nov 28, 2014, BIOCYC13B.