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Catalytic Site Atlas

CSA LITERATURE entry for 1uk7

E.C. name2-hydroxymuconate-semialdehyde hydrolase
SpeciesPseudomonas fluorescens (Bacteria)
E.C. Number (IntEnz)
CSA Homologues of 1uk7There are 28 Homologs
CSA Entries With UniProtID
CSA Entries With EC Number
PDBe Entry 1uk7
PDBSum Entry 1uk7
MACiE Entry 1uk7

Literature Report

IntroductionMeta-cleavage product hydrolases (MCP-hydrolase) are key enzymes involved in the microbial degradation of aromatic compounds. MCP-hydrolase CumD is a dimeric enzyme produced by Pseudomonas fluorescens IP01 and is involved in the pathway for the degradation of cumene. MCP-hydrolase enzymes produce 2-hydroxypenta-2,4-dienoate and various organic acids, depending upon the C6 substituent of the substrate. CumD prefers larger C6 substituents compared to other monoalkylbenzene hydrolases, such as TodF. Hence, when CumD acts on the meta-cleavage product of cumene, 2-hydroxypenta-2,4-dienoate and isobutyric acid are the products.
The understanding of the catalytic mechanism of MCP-hydrolases is neccessary to improve biological degradation of environmental pollutant aromatic compounds, which may be carcinogenic, toxic and mutagenic.
MechansimThe active site of CumD contains the Ser103, Asp 224, His 252 catalytic triad. Studies have shown that reaction proceeds not through initial nucleophilic attack, but via base catalysed attack on water by Ser 103.
1. The His 252 deprotonates Ser 103, activating it to basic attack of a water molecule. 2. The Ser 103 deprotonates the water, which then attacks the carbonyl carbon of the substrate. 3. The His and Ser residues form an active site ion pair, which is stabilised by the Asp 224 residue. 4. Hydrolysis of the substrate occurs, cleaving the C-C bond

Catalytic Sites for 1uk7

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
AlaA103103macie:sideChainSer 103 is deprotonated by his 252, and so can then act as a general base on a water molecule, activating it towards nucleophilic attack of the substrate carbonyl carbon. The negatively charged side chain Ser oxygen and the His imidazole ring form an active site ion pair.
AspA224224macie:sideChainThe Asp 224 forms a hydrogen bond to the His 252 to stabilise the ion pair.
HisA252252macie:sideChainThe his 252 deprotonates an adjacent serine residue and also forms an active site ion pair with the charged serine to stabilise the transition state.

Literature References

Notes:The Pseudomonas fluorescens species is able to grow on cumene and toluene as the sole source of carbon, but it cannot grow on biphenyl because the meta-cleavage pathway is blocked at the CumD step. Ile and Trp 143 on the helix alpha-4 appear to cause steric hinderance with the aromatic ring of the substrate, hampering base-catalysed attack by water.
Fushinobu S
A series of crystal structures of a meta-cleavage product hydrolase from Pseudomonas fluorescens IP01 (CumD) complexed with various cleavage products.
Biosci Biotechnol Biochem 2005 69 491-498
PubMed: 15784976
Saku T
Purification, characterization, and steady-state kinetics of a meta-cleavage compound hydrolase from Pseudomonas fluorescens IPO1.
J Biosci Bioeng 2002 93 568-574
PubMed: 16233251