Recombinant Peromyscus mexicanus NADH-ubiquinone oxidoreductase chain 3 (MT-ND3)
Description
Introduction to MT-ND3 Protein
NADH-ubiquinone oxidoreductase chain 3 (MT-ND3) is an essential component of Complex I in the mitochondrial respiratory chain. This protein plays a crucial role in the first step of electron transport during oxidative phosphorylation, the primary cellular process for ATP production. The MT-ND3 gene is encoded in the mitochondrial genome and is known by several synonyms including MTND3, NADH3, and ND3 . As a subunit of NADH dehydrogenase (EC 1.6.5.3), it participates in the transfer of electrons from NADH to ubiquinone within the inner mitochondrial membrane .
The mitochondrial location of the MT-ND3 gene makes it particularly valuable for evolutionary studies. Mitochondrial DNA (mtDNA) has become a powerful tool for assessing relationships among individuals, populations, and species due to its maternal inheritance pattern and relatively rapid mutation rate . This characteristic has made MT-ND3 and other mitochondrial genes important markers for phylogenetic analyses in diverse taxonomic groups, including Peromyscus rodents.
Taxonomic Context of Peromyscus mexicanus
Peromyscus mexicanus, commonly known as the Mexican deer mouse, is an endemic species found in southern Mexico with several recognized subspecies including P. m. teapensis, P. m. tehuantepecus, and P. m. totontepecus . The taxonomic understanding of this species has evolved significantly through molecular studies, particularly those examining mitochondrial DNA.
Recent mitochondrial DNA research has revealed that P. mexicanus exhibits a paraphyletic relationship pattern, meaning some of its subspecies are more closely related to other Peromyscus species than to other P. mexicanus subspecies. Specifically, molecular evidence indicates that P. m. teapensis, P. m. tehuantepecus, and P. m. totontepecus are more closely related to P. gymnotis than to other P. mexicanus populations . This finding challenges traditional morphology-based classifications and highlights the complex evolutionary history of this group.
The heterogeneous topography of Southern Mexico has contributed significantly to the diverse small mammal fauna in the region, leading to numerous complex taxonomic arrangements that continue to be refined through molecular studies . Mitochondrial genes like MT-ND3 have been instrumental in clarifying these relationships and understanding speciation patterns in this geographically complex region.
Structure and Characteristics of MT-ND3
Based on data from related Peromyscus species, the MT-ND3 protein from P. mexicanus is expected to be a relatively small protein of approximately 115 amino acids . The protein likely contains multiple hydrophobic domains consistent with its role as a membrane-embedded component of the respiratory chain complex I.
The amino acid sequences of MT-ND3 from related Peromyscus species show significant conservation with some species-specific variations. For comparative analysis, the known sequences from closely related species are presented below:
| Species | Amino Acid Sequence | Length |
|---|---|---|
| P. sejugis | MNMLTALLVNITLSMLLIIVAFWFFQLNLYTEKANPYECGFDPMGSARLPFSMKFFLVAI TFLLFDLEIALLLPLPWAIQMYNTNIMMLTAFILISVLALGLAYEWLQKGLEWTE | 115 aa |
| P. melanotis | MNMLMALLVNITLSTLLIIVAFWLPQLNLYTEKANPYECGFDPMGSARLPFSMKFFLVAI TFLLFDLEIALLLPLPWAIQMYNINIMMLTAFILVSVLALGLAYEWVQKGLEWTE | 115 aa |
| P. maniculatus | MNMLTALLVNITLSMLLIIVAFWLPQLNLYTEKANPYECGFDPMGSARLPFSMKFFLVAI TFLLFDLEIALLLPLPWAIQMYNIDIMMmLTAFILVSVLALGLAYEWLQKGLEWTE | 115 aa |
These sequences exhibit high conservation, particularly in functional domains essential for electron transport . While the exact sequence for P. mexicanus MT-ND3 is not provided in the available search results, it would likely show similar conservation patterns with minor variations reflecting its evolutionary relationship within the Peromyscus genus.
Production of Recombinant MT-ND3
The recombinant production of MT-ND3 protein from Peromyscus species typically employs bacterial expression systems, particularly Escherichia coli. Based on procedures for related species, the production of recombinant P. mexicanus MT-ND3 would likely follow a similar protocol.
Expression and Purification
The production process typically involves cloning the MT-ND3 gene into an expression vector with a His-tag sequence (usually N-terminal) to facilitate purification . After transformation into E. coli, protein expression is induced, followed by cell harvesting and lysis. The recombinant protein is then purified using affinity chromatography, taking advantage of the His-tag's affinity for metal ions.
The purified protein undergoes quality control assessment, typically using SDS-PAGE to confirm purity levels greater than 90% . The final product is usually supplied as a lyophilized powder to ensure stability during shipping and storage.
Protein Characteristics
The recombinant MT-ND3 from P. mexicanus would likely have the following characteristics, based on related Peromyscus species:
| Feature | Description |
|---|---|
| Expression System | E. coli |
| Protein Tag | His-tag (likely N-terminal) |
| Form | Lyophilized powder |
| Purity | >90% (SDS-PAGE) |
| Storage Buffer | Tris/PBS-based buffer, 6% Trehalose, pH 8.0 |
For reconstitution, it's typically recommended to briefly centrifuge the vial before opening to bring the contents to the bottom and then reconstitute the protein in deionized sterile water to a concentration of 0.1-1.0 mg/mL . Addition of glycerol (5-50% final concentration) is recommended for long-term storage.
Research Applications
Recombinant MT-ND3 from P. mexicanus has several important research applications spanning from basic biochemistry to evolutionary biology.
Mitochondrial Function Studies
As a component of respiratory chain Complex I, recombinant MT-ND3 is valuable for investigating mitochondrial function and energy metabolism. Researchers can use the purified protein to study electron transport mechanisms, complex assembly, and interactions with other mitochondrial components.
Phylogenetic and Evolutionary Studies
Perhaps the most significant application of P. mexicanus MT-ND3 is in evolutionary biology. Mitochondrial genes have become powerful markers for resolving taxonomic relationships due to their maternal inheritance and relatively rapid evolutionary rate. The paraphyletic nature of P. mexicanus revealed through mitochondrial DNA studies highlights the value of these markers in clarifying complex taxonomic relationships . Studies using MT-ND3 and other mitochondrial genes have challenged traditional morphology-based classifications and provided new insights into the evolutionary history of Peromyscus in Mexico.
Antibody Production and Protein Interaction Studies
Recombinant MT-ND3 can serve as an antigen for generating specific antibodies, which are valuable tools for protein localization, expression analysis, and immunoprecipitation studies. These antibodies can help researchers investigate the expression patterns of MT-ND3 in different tissues or under various physiological conditions.
Comparative Biochemistry
The availability of recombinant MT-ND3 from different Peromyscus species enables comparative biochemical studies to investigate how structural variations correlate with functional differences. Such studies can provide insights into the molecular basis of adaptation to different ecological niches.
Comparative Analysis with Related Species
The relationship between P. mexicanus and other Peromyscus species provides an interesting framework for comparative analysis. Recent molecular studies have revealed complex phylogenetic relationships that impact our understanding of MT-ND3 evolution in this genus.
Molecular Phylogeny
Mitochondrial DNA studies have shown that P. mexicanus is paraphyletic, with some subspecies (P. m. teapensis, P. m. tehuantepecus, and P. m. totontepecus) more closely related to P. gymnotis than to other P. mexicanus populations . This finding has significant implications for understanding the evolution of mitochondrial genes like MT-ND3 in this group.
Protein Comparison
Comparison of MT-ND3 proteins from different Peromyscus species reveals patterns of conservation and divergence that reflect their evolutionary relationships:
| Species | UniProt ID | Key Features | Ecological Distribution |
|---|---|---|---|
| P. sejugis | Q96114 | 115 aa, His-tagged recombinant | Santa Cruz mouse, endemic to Mexico |
| P. melanotis | Q95897 | 115 aa, His-tagged recombinant | Black-eared mouse, found in Mexico |
| P. maniculatus | Q95891 | 115 aa, recombinant protein | North American deer mouse, widespread |
| P. mexicanus | - | Expected 115 aa (inferred) | Mexican deer mouse, Southern Mexico |
The sequence similarities between these species reflect their close evolutionary relationships, while the differences may represent adaptations to different ecological conditions or genetic drift over evolutionary time .
Future Research Directions
Research on recombinant P. mexicanus MT-ND3 has several promising future directions:
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Complete sequence determination and structural characterization of P. mexicanus MT-ND3 to understand species-specific features
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Functional studies comparing MT-ND3 from different Peromyscus species to investigate potential adaptations in mitochondrial function
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Investigation of MT-ND3 variations across P. mexicanus subspecies to further clarify the paraphyletic relationships revealed by broader mitochondrial studies
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Development of specific antibodies against P. mexicanus MT-ND3 for expression studies in different tissues and developmental stages
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Exploration of potential applications in biomedical research, particularly in relation to mitochondrial disorders associated with MT-ND3 mutations
These research directions would contribute significantly to our understanding of mitochondrial biology, evolution, and potentially have applications in conservation biology for this endemic Mexican species.
Product Specs
Note: We will prioritize shipping the format we have in stock. However, if you have any specific format requirements, please indicate them when placing your order. We will prepare the product according to your request.
Note: All our proteins are shipped with standard blue ice packs. If you require dry ice shipping, please communicate with us in advance, as additional fees will apply.
Generally, the shelf life of liquid form is 6 months at -20°C/-80°C. The shelf life of lyophilized form is 12 months at -20°C/-80°C.
Target Background
Q&A
Basic Research Questions
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What is NADH-ubiquinone oxidoreductase chain 3 (MT-ND3) and what is its function in Peromyscus mexicanus?
MT-ND3 is a core subunit of the mitochondrial membrane respiratory chain NADH dehydrogenase (Complex I) that belongs to the minimal assembly required for catalysis. In Peromyscus mexicanus, this protein is 115 amino acids long with a molecular mass of approximately 13.3 kDa. It functions primarily in the transfer of electrons from NADH to the respiratory chain, with ubiquinone serving as the immediate electron acceptor for the enzyme. This electron transfer process is essential for oxidative phosphorylation and subsequent ATP production in mitochondria .
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What are the structural characteristics of Peromyscus mexicanus MT-ND3?
Peromyscus mexicanus MT-ND3 consists of 115 amino acids with the following sequence: MNMLMVMLVNITLSSCLIMIAFWLPQLNLYTEKANPYECGFDPMSSARLPFSMKFFLVAITFFLFDLEIALLLPLPWAIQMYNINTMMLTAFILVSVLALGLAYEWMQKGLEWTE . With a molecular mass of 13.3 kDa, this highly hydrophobic protein contains multiple transmembrane domains that anchor it within the inner mitochondrial membrane. As a member of the complex I subunit 3 family, MT-ND3's structure reflects its evolutionary conservation and essential role in energy metabolism across mammalian species.
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How does MT-ND3 contribute to mitochondrial function in Peromyscus species?
MT-ND3 plays a critical role in electron transport chain functionality through its participation in Complex I assembly and activity. Research has demonstrated that variants in MT-ND3 can significantly lower protein levels, causing Complex I assembly deficiencies, reduced activity, and diminished ATP synthesis . This direct relationship between MT-ND3 integrity and ATP production highlights its essential contribution to oxidative phosphorylation in Peromyscus mitochondria. The protein's evolutionary conservation across rodent species further underscores its fundamental importance in maintaining proper mitochondrial function.
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What methods are typically used to express and purify recombinant Peromyscus mexicanus MT-ND3?
The expression of recombinant MT-ND3 typically involves codon optimization for nuclear expression followed by translation via cytoplasmic ribosomes. This approach requires careful construction of mitochondrial targeting sequences along with the codon-optimized MT-ND3 gene to ensure proper import into the mitochondria . Purification often employs affinity chromatography techniques, particularly for His-tagged versions of the protein . Due to MT-ND3's hydrophobic nature, specialized detergents and buffer systems are necessary during purification to maintain protein stability and native conformation. The expression system selection is critical, with both bacterial and mammalian systems offering distinct advantages depending on the experimental requirements.
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What are the common applications of recombinant Peromyscus MT-ND3 in research?
Recombinant Peromyscus MT-ND3 serves multiple research purposes, including:
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Investigation of Complex I assembly and activity in normal and pathological conditions
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Functional characterization of disease-associated MT-ND3 variants
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Complementation studies to rescue mitochondrial dysfunction
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Comparative analyses across Peromyscus species to study evolutionary conservation
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Development of therapeutic approaches for mitochondrial disorders
Additionally, as Peromyscus represents an important mammalian research model distinct from traditional Mus and Rattus systems, studies involving its mitochondrial components like MT-ND3 offer unique insights into physiological adaptations and disease mechanisms .
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