Recombinant Mouse Enoyl-CoA hydratase, mitochondrial (Echs1) is produced in E. coli and covers the full length of the mature protein from amino acids 28 to 290 with a C225S mutation. The protein features an N-terminal 10xHis tag and a C-terminal HA tag, which should make purification and detection more straightforward. SDS-PAGE analysis indicates it reaches a purity level greater than 85%, suggesting it's well-suited for various research applications.
Enoyl-CoA hydratase, mitochondrial (Echs1) appears to play a critical role in the fatty acid oxidation pathway. It catalyzes the hydration of enoyl-CoA to hydroxyacyl-CoA—a step that seems essential for breaking down fatty acids and extracting energy from them. This makes Echs1 particularly relevant for studies examining metabolic processes and energy homeostasis, potentially offering valuable insights into how cells manage their metabolism.
Potential Applications
Note: The applications listed below are based on what we know about this protein's biological functions, published research, and experience from experts in the field. However, we haven't fully tested all of these applications ourselves yet. We'd recommend running some preliminary tests first to make sure they work for your specific research goals.
1. Biochemical Characterization of Mutant Enzyme Activity
This recombinant Echs1 protein carries a C225S mutation that may help researchers understand what role cysteine-225 plays in enzyme function and stability. Comparative kinetic assays between wild-type and mutant forms could reveal how much this specific residue contributes to catalytic activity. The dual His and HA tags should simplify both purification and detection, making detailed biochemical analysis more feasible—including substrate binding studies and thermal stability measurements.
2. Protein-Protein Interaction Studies
Both the N-terminal His tag and C-terminal HA tag make this protein a good candidate for pull-down assays aimed at identifying potential binding partners of Echs1 in mitochondrial fatty acid oxidation pathways. These tags can be put to work in co-immunoprecipitation experiments or affinity chromatography to isolate protein complexes. Such an approach might help clarify the molecular interactions involved in mitochondrial β-oxidation and metabolic regulation, though the complexity of these pathways suggests results may require careful interpretation.
3. Antibody Development and Validation
The recombinant protein can serve as an antigen for generating specific antibodies against mouse Echs1. This appears particularly useful for researchers studying mitochondrial metabolism in mouse models. The HA tag provides an internal control for immunoassays, while the high purity (>85%) should ensure reliable antibody production. Generated antibodies can then be validated using this recombinant protein in Western blot, ELISA, and immunofluorescence applications.
4. Structural and Folding Studies
The C225S mutation presents an opportunity to investigate how amino acid substitutions might affect protein folding and structural integrity of the enoyl-CoA hydratase domain. Researchers could use this mutant protein for comparative structural studies through techniques like circular dichroism spectroscopy, dynamic light scattering, or crystallography. The dual tags help with protein tracking during purification and concentration steps that structural analysis typically requires.
5. In Vitro Metabolic Pathway Reconstitution
This recombinant Echs1 mutant can be incorporated into in vitro fatty acid oxidation assays to examine how the C225S mutation affects overall pathway efficiency. Researchers might combine this protein with other purified enzymes from the β-oxidation pathway to reconstruct the complete metabolic process under controlled conditions. The tags allow for easy monitoring of protein levels and stability during the extended incubation periods that metabolic studies often demand.
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