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Escherichia coli K-12 substr. MG1655 Enzyme: 2-amino-3-ketobutyrate CoA ligase



Gene: kbl Accession Numbers: EG10512 (EcoCyc), b3617, ECK3607

Regulation Summary Diagram: ?

Subunit composition of 2-amino-3-ketobutyrate CoA ligase = [Kbl]2

Summary:
2-Amino-3-ketobutyrate CoA ligase (Kbl, AKB ligase) catalyzes the reversible, coenzyme A-dependent cleavage/condensation reaction between 2-amino-3-oxobutanoate (2-amino-3-ketobutyrate) and glycine plus acetyl-CoA. It is the second reaction in the threonine dehydrogenase-initiated pathway by which threonine is converted to glycine which is then converted to serine. This pathway is the primary route for threonine utilization in prokaryotes and is an alternate pathway for serine biosynthesis in E. coli [Mukherjee87, Ravnikar87]. Because 2-amino-3-oxobutanoate can spontaneously decarboxylate to aminoacetone, a second threonine dehydrogenase-initiated pathway is also possible (see threonine degradation III (to methylglyoxal)).

The crystal structure of Kbl in complex with a pyridoxal-5'-phosphate-substrate intermediate has been determined at 2.0 Å resolution and a reaction mechanism was proposed. Kbl belongs to the α family of PLP-dependent enzymes. It was noted that this enzyme has been evolutionarily conserved and E. coli Kbl shares 54% amino acid sequence identity with the human enzyme [Schmidt01]. Details of the condensation mechanism have been studied [Bashir06].

Review: Reitzer, L. (2005) "Catabolism of Amino Acids and Related Compounds" EcoSal 3.4.7 [ECOSAL]

Gene Citations: [Aronson88, Rex91, Landgraf94]

Locations: cytosol

Map Position: [3,789,378 <- 3,790,574] (81.67 centisomes)
Length: 1197 bp / 398 aa

Molecular Weight of Polypeptide: 43.117 kD (from nucleotide sequence), 42.0 kD (experimental) [Mukherjee87 ]

Molecular Weight of Multimer: 85.0 kD (experimental) [Mukherjee87]

pI: 5.99

Unification Links: ASAP:ABE-0011832 , CGSC:18205 , DIP:DIP-48030N , EchoBASE:EB0507 , EcoGene:EG10512 , EcoliWiki:b3617 , Mint:MINT-1315923 , ModBase:P0AB77 , OU-Microarray:b3617 , PortEco:kbl , PR:PRO_000023046 , Pride:P0AB77 , Protein Model Portal:P0AB77 , RefSeq:NP_418074 , RegulonDB:EG10512 , SMR:P0AB77 , String:511145.b3617 , UniProt:P0AB77

Relationship Links: InterPro:IN-FAMILY:IPR001917 , InterPro:IN-FAMILY:IPR004839 , InterPro:IN-FAMILY:IPR011282 , InterPro:IN-FAMILY:IPR015421 , InterPro:IN-FAMILY:IPR015422 , InterPro:IN-FAMILY:IPR015424 , PDB:Structure:1FC4 , Pfam:IN-FAMILY:PF00155 , Prosite:IN-FAMILY:PS00599

In Paralogous Gene Group: 193 (2 members)

Gene-Reaction Schematic: ?

Genetic Regulation Schematic: ?

GO Terms:

Biological Process: GO:0008152 - metabolic process Inferred by computational analysis [GOA01a]
GO:0009058 - biosynthetic process Inferred by computational analysis [GOA01a]
GO:0019518 - L-threonine catabolic process to glycine Inferred by computational analysis [UniProtGOA12]
Molecular Function: GO:0016874 - ligase activity Inferred from experiment [Mukherjee87]
GO:0030170 - pyridoxal phosphate binding Inferred from experiment Inferred by computational analysis [GOA01a, Mukherjee87, Schmidt01]
GO:0042803 - protein homodimerization activity Inferred from experiment [Mukherjee87]
GO:0046872 - metal ion binding Inferred from experiment [Mukherjee87]
GO:0003824 - catalytic activity Inferred by computational analysis [GOA01a]
GO:0008890 - glycine C-acetyltransferase activity Inferred by computational analysis [GOA01, GOA01a]
GO:0016740 - transferase activity Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0016746 - transferase activity, transferring acyl groups Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005737 - cytoplasm Inferred from experiment [Schmidt01]
GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, LopezCampistrou05]

MultiFun Terms: metabolism carbon utilization amino acids
metabolism central intermediary metabolism threonine catabolism

Essentiality data for kbl 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]
Yes [Feist07, Comment 4]

Credits:
Last-Curated ? 16-Nov-2011 by Fulcher C , SRI International


Enzymatic reaction of: 2-amino-3-ketobutyrate CoA ligase

Synonyms: glycine C-acetyltransferase, AKB ligase, aminoacetone synthase, aminoacetone synthetase, acetyl-CoA:glycine C-acetyltransferase

EC Number: 2.3.1.29

glycine + acetyl-CoA <=> 2-amino-3-oxobutanoate + coenzyme A + H+

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. [Mukherjee87]

Alternative Substrates for acetyl-CoA [Comment 5 ]: butanoyl-CoA [Mukherjee87 ] , propanoyl-CoA [Mukherjee87 ]

In Pathways: superpathway of threonine metabolism , threonine degradation II

Summary:
The enzyme activity was assayed in the reverse direction as the amount of coenzyme A released by the condensation of acetyl-CoA with glycine. No activity was seen with valeryl-CoA, succinyl-CoA, or glutaryl-CoA. The enzyme was specific for glycine and no activity was seen with glycinamide, glycine methylester, 2-aminoethanol, aminomethylphosphonic acid, aminomalonate, L-alanine, L-serine, L-threonine, L-valine, or L-leucine. Varying degrees of inhibition were seen with the divalent cations Mn2+, Ni2+ Ca2+ Sr2+ Ba2+ Zn2+ Hg2+ Cu2+ and Cd2+, with the latter three showing strongest inhibition [Mukherjee87].

The enzyme was also shown to have L-threonine aldolase activity, cleaving L-threonine to acetaldehyde and glycine. Although the Km for glycine (ligase activity) was 10 mM and the Km for L-threonine (aldolase activity) was 0.9 mM, the Vmax values were 2.5 μmol of CoA released/min per mg for the ligase activity versus 0.014 μmol of acetaldehyde formed (NADH oxidized)/min per mg for the aldolase activity [Marcus93].

Cofactors or Prosthetic Groups: pyridoxal 5'-phosphate [Comment 6]

Inhibitors (Competitive): aminomalonate [Comment 7]

Inhibitors (Unknown Mechanism): diacetyl [Mukherjee92] , cyclohexane-1,2-dione [Mukherjee92] , phenylglyoxal [Mukherjee92] , 4-(oxoacetyl)phenoxyacetate [Mukherjee92] , Hg2+ [Mukherjee87] , Cu2+ [Mukherjee87] , Cd2+ [Mukherjee87] , L-cysteine [Mukherjee87] , dithiothreitol [Mukherjee87] , glutathione [Mukherjee87] , aminomethylphosphonate [Mukherjee87] , coenzyme A [Comment 8]

Primary Physiological Regulators of Enzyme Activity: L-cysteine , glutathione , coenzyme A , aminomalonate

Kinetic Parameters:

Substrate
Km (μM)
Citations
acetyl-CoA
59.0
[Mukherjee87, BRENDA14]
glycine
12000.0
[Mukherjee87]

pH(opt): 7.5 [BRENDA14, Mukherjee87]


Sequence Features

Feature Class Location Citations Comment
Sequence-Conflict 43
[Aronson88, UniProt10a]
Alternate sequence: H → Q; UniProt: (in Ref. 1; CAA29883);
Protein-Segment 111 -> 112
[UniProt10]
UniProt: Pyridoxal phosphate binding; Sequence Annotation Type: region of interest;
Amino-Acid-Sites-That-Bind 136
[UniProt10a]
UniProt: Pyridoxal phosphate;
Sequence-Conflict 171
[Aronson88, UniProt10a]
Alternate sequence: A → R; UniProt: (in Ref. 1; CAA29883);
Sequence-Conflict 183
[Aronson88, UniProt10a]
Alternate sequence: V → L; UniProt: (in Ref. 1; CAA29883);
Amino-Acid-Sites-That-Bind 185
[UniProt10a]
UniProt: Pyridoxal phosphate;
Protein-Segment 210 -> 213
[UniProt10]
UniProt: Pyridoxal phosphate binding; Sequence Annotation Type: region of interest;
Amino-Acid-Sites-That-Bind 213
[UniProt10a]
UniProt: Substrate;
Protein-Segment 241 -> 244
[UniProt10]
UniProt: Pyridoxal phosphate binding; Sequence Annotation Type: region of interest;
N6-pyridoxal-phosphate-Lys-Modification 244
[UniProt11a]
UniProt: N6-(pyridoxal phosphate)lysine.
Amino-Acid-Sites-That-Bind 244
[UniProt10a]
UniProt: Substrate;
Protein-Segment 274 -> 275
[UniProt10]
UniProt: Pyridoxal phosphate binding; Sequence Annotation Type: region of interest;
Amino-Acid-Sites-That-Bind 368
[UniProt10a]
UniProt: Substrate;


Gene Local Context (not to scale): ?

Transcription Unit:

Notes:

History:
10/20/97 Gene b3617 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10512.


References

Aronson88: Aronson BD, Ravnikar PD, Somerville RL (1988). "Nucleotide sequence of the 2-amino-3-ketobutyrate coenzyme A ligase (kbl) gene of E. coli." Nucleic Acids Res 16(8);3586. PMID: 3287333

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

Bashir06: Bashir Q, Rashid N, Akhtar M (2006). "Mechanism and substrate stereochemistry of 2-amino-3-oxobutyrate CoA ligase: implications for 5-aminolevulinate synthase and related enzymes." Chem Commun (Camb) (48);5065-7. PMID: 17146529

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

ECOSAL: "Escherichia coli and Salmonella: Cellular and Molecular Biology." Online edition.

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."

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

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

Landgraf94: Landgraf JR, Levinthal M, Danchin A (1994). "The role of H-NS in one carbon metabolism." Biochimie 1994;76(10-11);1063-70. PMID: 7748928

LopezCampistrou05: Lopez-Campistrous A, Semchuk P, Burke L, Palmer-Stone T, Brokx SJ, Broderick G, Bottorff D, Bolch S, Weiner JH, Ellison MJ (2005). "Localization, annotation, and comparison of the Escherichia coli K-12 proteome under two states of growth." Mol Cell Proteomics 4(8);1205-9. PMID: 15911532

Marcus93: Marcus JP, Dekker EE (1993). "Identity and some properties of the L-threonine aldolase activity manifested by pure 2-amino-3-ketobutyrate ligase of Escherichia coli." Biochim Biophys Acta 1164(3);299-304. PMID: 8343529

Mukherjee87: Mukherjee JJ, Dekker EE (1987). "Purification, properties, and N-terminal amino acid sequence of homogeneous Escherichia coli 2-amino-3-ketobutyrate CoA ligase, a pyridoxal phosphate-dependent enzyme." J Biol Chem 1987;262(30);14441-7. PMID: 3117785

Mukherjee90: Mukherjee JJ, Dekker EE (1990). "2-Amino-3-ketobutyrate CoA ligase of Escherichia coli: stoichiometry of pyridoxal phosphate binding and location of the pyridoxyllysine peptide in the primary structure of the enzyme." Biochim Biophys Acta 1990;1037(1);24-9. PMID: 2104756

Mukherjee92: Mukherjee JJ, Dekker EE (1992). "Inactivation of Escherichia coli 2-amino-3-ketobutyrate CoA ligase by phenylglyoxal and identification of an active-site arginine peptide." Arch Biochem Biophys 299(1);147-53. PMID: 1444446

Ravnikar87: Ravnikar PD, Somerville RL (1987). "Genetic characterization of a highly efficient alternate pathway of serine biosynthesis in Escherichia coli." J Bacteriol 169(6);2611-7. PMID: 3108237

Rex91: Rex JH, Aronson BD, Somerville RL (1991). "The tdh and serA operons of Escherichia coli: mutational analysis of the regulatory elements of leucine-responsive genes." J Bacteriol 1991;173(19);5944-53. PMID: 1917830

Schmidt01: Schmidt A, Sivaraman J, Li Y, Larocque R, Barbosa JA, Smith C, Matte A, Schrag JD, Cygler M (2001). "Three-dimensional structure of 2-amino-3-ketobutyrate CoA ligase from Escherichia coli complexed with a PLP-substrate intermediate: inferred reaction mechanism." Biochemistry 40(17);5151-60. PMID: 11318637

UniProt10: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

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

UniProt11a: UniProt Consortium (2011). "UniProt version 2011-11 released on 2011-11-22 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

Fraenkel95: Fraenkel YM, Mandel Y, Friedberg D, Margalit H (1995). "Identification of common motifs in unaligned DNA sequences: application to Escherichia coli Lrp regulon." Comput Appl Biosci 1995;11(4);379-87. PMID: 8521047


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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 Sat Nov 22, 2014, biocyc14.