Recombinant Serpentine receptor class delta-51 (srd-51)
Description
Introduction to Serpentine Receptors in C. elegans
The nematode Caenorhabditis elegans possesses an extraordinarily diverse array of serpentine receptors that function as critical mediators in cellular signaling pathways. These receptors, characterized by their distinctive seven-transmembrane domain structure, constitute one of the largest protein families in the C. elegans genome, reflecting their evolutionary significance in chemosensation, development, and environmental adaptation. Serpentine receptors in C. elegans are categorized into several classes, including srd (serpentine receptor class delta), which encompasses SRD-51, the focus of this article.
The serpentine receptor classification system in C. elegans includes numerous distinct families, with many members being protein-coding genes. The genome database reveals multiple serpentine receptor variants, including srd-50 (Serpentine receptor class delta-50), srx-44 (Serpentine Receptor, class X), srx-51 (Serpentine Receptor, class X), and numerous others that participate in various physiological processes . Within this diverse receptor landscape, SRD-51 emerges as a notable protein-coding gene whose recombinant form has become valuable for scientific investigation.
Protein Structure and Sequence Analysis
SRD-51 is a full-length protein consisting of 336 amino acids derived from Caenorhabditis elegans. The amino acid sequence of SRD-51 reveals characteristic features typical of serpentine receptors, including hydrophobic transmembrane domains and functional motifs essential for signal transduction . The complete amino acid sequence of SRD-51 is:
MSEVEKKLEMFVTVYYSLNVTLALSINILLLFIMKTTKSSLLKDMQYYLFNTALFEIIVSLSTYFAQCRPVANKSTLAVFCHGPCKYFGKNTCFVTFAVVQCSVVAASFSILLSFYYRYRLKVNFKKKHKHATTFIIFSFFPTVMLLFQLLTDSNFAIVEAETREMHPDYDYVNNALIGFSDSKSPAAIIAQSLISLGVYMSPLIAFHYRRKINKILSTNTGQRIPVAYCKQLINGLLIIQTLIPFCVYIPPYSYFLYSQLSGHSNLYFEYLLNIFGSFTAFINPLLTFYFVLPYRRALCKKVFKYFPSISEEGTEITTFPTTVQFQRGHTASTKF
Analysis of this sequence reveals typical structural elements of serpentine receptors, including hydrophobic transmembrane domains, extracellular and intracellular loops, and potential ligand-binding regions. The presence of conserved cysteine residues suggests the formation of disulfide bonds that may stabilize the tertiary structure.
Functional Domains and Motifs
Examination of the SRD-51 sequence reveals several noteworthy features:
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Signal peptide region at the N-terminus (approximately residues 1-20)
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Multiple transmembrane domains characterized by hydrophobic amino acid stretches
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Potential ligand-binding pocket formed by extracellular loops
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Intracellular regions likely involved in G-protein coupling and downstream signaling
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Conserved motifs typical of class delta serpentine receptors
These structural elements collectively contribute to the receptor's presumed function in signal transduction, though specific ligands and signaling pathways for SRD-51 remain areas for further investigation.
Expression Systems and Methodology
The recombinant form of SRD-51 has been successfully produced using bacterial expression systems, specifically Escherichia coli. This approach allows for scalable production of the protein for research applications. The recombinant SRD-51 is typically engineered with an N-terminal histidine tag (His-tag), facilitating purification through affinity chromatography and potentially enhancing solubility .
The production process involves several key steps:
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Cloning of the srd-51 gene sequence into an appropriate expression vector
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Transformation of E. coli host cells with the recombinant construct
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Induction of protein expression under controlled conditions
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Cell harvest and lysis to release the recombinant protein
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Purification using affinity chromatography targeting the His-tag
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Additional purification steps to enhance purity (>90% as determined by SDS-PAGE)
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Lyophilization to produce a stable powder form for storage
Functional Characterization and Signaling Pathways
While specific functions of SRD-51 remain to be fully elucidated, recombinant protein can facilitate investigations into potential signaling pathways. Based on homology with other serpentine receptors, SRD-51 likely couples to G proteins to initiate intracellular signaling cascades in response to specific ligands, potentially mediating chemosensory functions or other physiological processes in C. elegans.
Comparative Analysis with Related Receptors
The C. elegans genome contains numerous serpentine receptors with varying degrees of sequence similarity to SRD-51. Comparative analysis of recombinant SRD-51 with related receptors such as SRD-50 could reveal evolutionary relationships and functional divergence within this receptor family . Such comparisons may provide insights into the specialization of different receptor classes for distinct physiological roles.
Genomic Organization and Expression
The srd-51 gene belongs to the broader landscape of protein-coding genes in C. elegans. The nematode genome contains numerous serpentine receptor genes organized into distinct classes, reflecting the evolutionary importance of these receptors for the organism's sensory capabilities and environmental interactions .
Understanding the genomic context of srd-51 requires consideration of its relationship to other serpentine receptor genes, including potential gene duplications, conserved regulatory elements, and expression patterns across different tissues and developmental stages.
Evolutionary Conservation and Divergence
Comparative genomic analyses could reveal the evolutionary history of srd-51 and related genes. While specific information about SRD-51 conservation across species is limited in the provided search results, serpentine receptors generally show complex patterns of evolutionary dynamics, with evidence of lineage-specific expansions and functional divergence.
The availability of recombinant SRD-51 facilitates phylogenetic studies through sequence and structural comparisons with homologous proteins from other nematodes and more distantly related organisms.
Challenges in Membrane Protein Research
As a seven-transmembrane domain protein, SRD-51 presents inherent challenges for structural and functional studies due to its hydrophobic nature and dependence on membrane environment for proper folding. The recombinant production in E. coli may not fully recapitulate the native conformational state, potentially affecting functional assays.
Researchers utilizing recombinant SRD-51 should consider:
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Potential differences between bacterially-expressed protein and native form
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The impact of the His-tag on protein structure and function
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Challenges in reconstituting membrane proteins into appropriate lipid environments
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Limitations in mimicking the native cellular context for functional studies
Product Specs
Note: While we prioritize shipping the format currently in stock, please specify your format preference in order notes for customized fulfillment.
Note: All proteins are shipped with standard blue ice packs. Dry ice shipping requires advance notice and incurs additional charges.
The tag type is finalized during production. If you require a specific tag, please inform us, and we will prioritize its development.
Target Background
KEGG: cel:CELE_F15A2.3
UniGene: Cel.29422
Q&A
What is Serpentine receptor class delta-51 (srd-51) and what organism does it originate from?
Serpentine receptor class delta-51 (srd-51) is a G-protein coupled receptor (GPCR) belonging to the serpentine receptor class D family in Caenorhabditis elegans. It is encoded by the srd-51 gene, also known by its ORF name F15A2.3 . C. elegans is a free-living, transparent nematode that serves as an important model organism in developmental biology, neuroscience, and genetics research. The srd-51 protein is part of the broader family of serpentine receptors which are characterized by their transmembrane structure with seven membrane-spanning domains.
How does srd-51 compare to other serpentine receptors in C. elegans?
C. elegans contains a large and diverse family of serpentine receptors, with the serpentine receptor class D (srd) representing one subfamily. Within the nematode genome, there are numerous serpentine receptor classes including srh, str, sri, and srj, among others. These receptors are primarily expressed in chemosensory neurons and are thought to mediate responses to environmental chemicals, pheromones, and other stimuli.
The srd family belongs to the larger delta class of serpentine receptors in C. elegans . While specific comparative data for srd-51 is limited in the search results, serpentine receptors generally display significant sequence diversity in their ligand-binding domains while maintaining structural conservation in their transmembrane regions.
What phenotypes are associated with srd-51 mutations or knockdowns?
For researchers interested in identifying potential phenotypes associated with srd-51, similar bioinformatics approaches combined with RNAi validation could be employed. The search results indicate that such bioinformatics-based phenotype prediction has success rates of approximately 22% for sterility and 41% for uncoordinated phenotypes .
What experimental approaches are optimal for studying srd-51 function?
Several experimental approaches can be employed to study srd-51 function:
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RNA Interference (RNAi): RNAi can be used to selectively silence srd-51 gene expression and observe resulting phenotypic changes. The search results mention that RNAi has been successfully used to study genes in C. elegans, with subsequent validation through RT-PCR .
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Recombinant Protein Expression: Expression of recombinant srd-51 can facilitate structural studies and protein-protein interaction analyses. The recombinant protein should be expressed with appropriate tags for purification and detection .
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CRISPR-Cas9 Genome Editing: Creating precise deletions, insertions, or point mutations in the srd-51 gene can provide insights into the functional domains of the receptor.
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Transgenic Reporter Strains: Creating transgenic C. elegans strains expressing fluorescent proteins under the control of the srd-51 promoter can reveal expression patterns and temporal regulation.
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Calcium Imaging: Since serpentine receptors often mediate changes in intracellular calcium, calcium imaging in neurons expressing srd-51 during exposure to potential ligands can help identify receptor activators.
How can bioinformatics approaches enhance srd-51 research?
Bioinformatics approaches offer powerful tools for studying srd-51:
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Sequence Similarity Analysis: As demonstrated in Batra's research, sequence similarity analysis can predict phenotypes associated with specific genes with relatively high success rates .
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Structural Modeling: Using homology modeling based on known GPCR structures to predict the three-dimensional structure of srd-51.
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Ligand Prediction: Computational docking studies can predict potential ligands for srd-51 based on its predicted binding pocket.
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Expression Pattern Analysis: Mining existing RNA-seq and other transcriptomic datasets to identify conditions under which srd-51 is differentially expressed.
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Protein-Protein Interaction Networks: Predicting potential interaction partners based on known interactions of similar receptors.
What are the optimal conditions for working with recombinant srd-51 protein?
Based on the product information provided in the search results, recombinant srd-51 protein should be handled with the following considerations:
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Storage: Store at -20°C, with extended storage at -20°C or -80°C. Working aliquots can be stored at 4°C for up to one week .
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Buffer Conditions: The protein is typically maintained in a Tris-based buffer with 50% glycerol, optimized for stability .
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Handling Precautions: Repeated freezing and thawing should be avoided to maintain protein integrity .
When designing experiments using recombinant srd-51, researchers should consider these parameters to ensure optimal protein activity and stability.
How can RNA interference be effectively employed to study srd-51 function?
RNA interference is a powerful technique for studying gene function in C. elegans. For effective RNAi targeting srd-51:
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RNAi Design: Design double-stranded RNA (dsRNA) targeting unique regions of the srd-51 mRNA to avoid off-target effects.
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Delivery Methods: Several methods can be used:
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Feeding: Engineer E. coli to express dsRNA and feed these bacteria to worms
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Soaking: Immerse worms in dsRNA solution
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Injection: Directly inject dsRNA into the gonad
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Mechanism Understanding: RNAi in C. elegans involves:
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Validation: Confirm srd-51 knockdown using reverse transcriptase PCR (RT-PCR) to measure mRNA levels, as mentioned in the search results for other genes .
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Phenotypic Analysis: Carefully document any phenotypic changes resulting from srd-51 knockdown, looking for alterations in behavior, development, reproduction, or neural function.
What techniques are most effective for analyzing srd-51 protein interactions?
To analyze srd-51 protein interactions, researchers can employ several techniques:
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Co-immunoprecipitation (Co-IP): Using antibodies against srd-51 or its tagged version to pull down protein complexes, followed by mass spectrometry to identify interaction partners.
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Yeast Two-Hybrid (Y2H): Testing for direct protein-protein interactions by expressing srd-51 domains fused to DNA-binding domains and potential partners fused to activation domains.
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Bioluminescence Resonance Energy Transfer (BRET): Monitoring real-time protein-protein interactions in living cells by tagging srd-51 and candidate partners with appropriate donor and acceptor molecules.
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Surface Plasmon Resonance (SPR): Measuring binding kinetics between purified srd-51 and potential ligands or interaction partners.
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Proximity Labeling: Using techniques like BioID or APEX to identify proteins in close proximity to srd-51 in its native cellular environment.
How can integrative approaches advance our understanding of srd-51?
An integrative research approach combining multiple methodologies could significantly advance our understanding of srd-51:
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Combining Genomics and Phenomics: Integrate genome-wide association studies with phenotypic data to understand how genetic variations in srd-51 correlate with phenotypic changes.
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Multi-omics Integration: Combine transcriptomics, proteomics, and metabolomics data to understand how srd-51 functions within broader cellular networks.
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Comparative Studies: Explore srd-51 orthologs in other nematode species to understand evolutionary conservation and divergence of function.
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Translational Research: Investigate whether insights from srd-51 research in C. elegans have implications for understanding related receptors in other organisms, including humans.
What are the challenges in studying serpentine receptors like srd-51?
Researchers face several challenges when studying serpentine receptors like srd-51:
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Structural Determination: G-protein coupled receptors are notoriously difficult to crystallize for structural studies due to their transmembrane nature.
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Ligand Identification: Identifying the natural ligands for orphan receptors like srd-51 remains challenging.
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Functional Redundancy: C. elegans has many serpentine receptors with potentially overlapping functions, making it difficult to observe clear phenotypes from single gene manipulations.
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Expression Systems: Establishing expression systems that properly fold and traffic serpentine receptors while maintaining their functionality.
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Specificity of Tools: Developing specific antibodies and other molecular tools for srd-51 can be challenging due to sequence similarity with other serpentine receptors.
