Recombinant Phenylobacterium zucineum Urocanate hydratase (hutU), partial
Description
Overview of Urocanate Hydratase (hutU) in Bacterial Systems
Urocanate hydratase (EC 4.2.1.49) is a key enzyme in the histidine degradation pathway, catalyzing the conversion of urocanate to 4-imidazolone-5-propionate. While hutU is not explicitly annotated in the P. zucineum HLK1T genome , homologs of this enzyme are common in Alphaproteobacteria.
Functional Context:
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Role: Critical in nitrogen metabolism, enabling bacteria to utilize histidine as a carbon/nitrogen source.
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Structural Features: Typically requires Fe²⁺ or other divalent cations for activity.
Genomic Insights into P. zucineum Metabolic Pathways
The P. zucineum genome encodes enzymes for aromatic compound degradation (e.g., phenylalanine via the homogentisate pathway) but lacks explicit annotations for histidine catabolism . Key metabolic observations include:
Table 1: Key Metabolic Pathways in P. zucineum
Recombinant Enzyme Production Hypotheses
If P. zucineum hutU were to be recombinantly expressed, the following steps would apply based on its genomic features:
Cloning Strategy
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Gene Identification: Requires sequencing gaps to be resolved, as current annotations lack hutU .
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Vector Design: Plasmid pET or pGEX systems, leveraging P. zucineum’s codon usage bias (GC content: 71.2%) .
Table 2: Proposed Cloning Parameters
| Parameter | Value | Rationale |
|---|---|---|
| Host System | E. coli BL21(DE3) | High-yield cytoplasmic expression |
| Induction | IPTG (0.1–1 mM) | Compatible with T7/lac promoters |
| Tagging | His₆ or GST | Facilitates purification |
Challenges and Research Gaps
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Annotation Limitations: The HLK1T genome lacks explicit hutU annotation, suggesting either a pathway gap or horizontal gene transfer requirement .
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Functional Redundancy: Homologs in related species (e.g., Caulobacter crescentus) could guide heterologous expression studies.
Recommendations for Future Work
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Proteomic Screening: Use mass spectrometry to identify unannotated hydrolases in P. zucineum lysates.
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Comparative Genomics: Analyze Phenylobacterium species (e.g., P. lituiforme) for conserved hut operons.
Product Specs
Target Background
KEGG: pzu:PHZ_c0468
STRING: 450851.PHZ_c0468
