Recombinant Acinetobacter sp. Putative aminoacrylate hydrolase RutD (rutD)
Description
Introduction to Recombinant Acinetobacter sp. Putative Aminoacrylate Hydrolase RutD (rutD)
Recombinant Acinetobacter sp. putative aminoacrylate hydrolase RutD (rutD) is an enzyme that belongs to the α/β hydrolase superfamily and is involved in the rut pathway of pyrimidine degradation . The rut pathway enables organisms to use pyrimidine bases as a sole nitrogen source, particularly under suboptimal temperatures .
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Function: RutD is proposed to enhance the rate of hydrolysis of aminoacrylate, a toxic byproduct of uracil degradation, into malonic semialdehyde . Although this reaction can occur spontaneously in water, the toxicity of aminoacrylate necessitates catalysis by RutD for efficient growth when uracil is used as a nitrogen source .
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Characteristics: RutD exhibits a novel arrangement of residues in its active site, differing from the canonical catalytic triad found in other α/β hydrolases .
Catalytic Mechanism
RutD's catalytic triad consists of unusual composition, with histidine (H87) replacing the typical nucleophile found in other α/β hydrolases . This substitution is conserved among all known RutD proteins . A glycerol molecule was observed bound in the cleft formed by cap helices 6 and 7 and loops from the enzyme's core, occupying what is presumed to be an active site .
Role in Pyrimidine Catabolism
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Rut Pathway: The rut pathway consists of a pyrimidine transporter and six enzymes that cleave and process the uracil ring . The central enzymes, RutA and RutB, catalyze a reaction where the uracil ring is cleaved to produce ureidoacrylate . RutB then hydrolyzes ureidoacrylate into ammonium, malonic semialdehyde, and carbon dioxide . RutC and RutD neutralize the toxic effects of pathway intermediates .
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RutD's Specific Function: RutD facilitates the hydrolysis of aminoacrylate to malonic semialdehyde, which is crucial for neutralizing the toxicity of aminoacrylate during uracil degradation .
Biochemical Properties
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Purity: Recombinant RutD is typically >85% pure as determined by SDS-PAGE .
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Storage: Liquid form RutD can be stored for 6 months at -20°C/-80°C. Lyophilized form RutD can be stored for 12 months at -20°C/-80°C . Repeated freezing and thawing is not recommended .
Genetic Information
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Uniprot No.: The Uniprot accession number for RutD is Q6FFZ9 .
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Immunogen Species: Acinetobacter baylyi (strain ATCC 33305 / BD413 / ADP1) .
Active Site Residues Conservation
| Residues Forming Active Site | % of Conservancy |
|---|---|
| G21 | 100 |
| L22 | 99 |
| G23 | 100 |
| G24 | 100 |
| H87 | 100 |
| A88 | 88 |
| R128 | 100 |
| Q144 | 88 |
| L148 | 91 |
| D209 | 100 |
| L211 | 91 |
| H237 | 100 |
Comparison with Other Hydrolases
| Protein Name | RutD | PcaD |
|---|---|---|
| Nucleophile | H87 | S100 |
| Acidic Residue | D209 | D217 |
| Histidine | H237 | H244 |
Product Specs
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Target Background
KEGG: aci:ACIAD0025
STRING: 62977.ACIAD0025
