Recombinant Bacillus pumilus UPF0295 protein BPUM_0828 (BPUM_0828)

What is Bacillus pumilus UPF0295 protein BPUM_0828?

Bacillus pumilus UPF0295 protein BPUM_0828 is a full-length protein consisting of 115 amino acids with UniProt ID A8FB95. The protein belongs to the UPF0295 family, which contains proteins with currently undefined functions that require further characterization . The complete amino acid sequence of this protein is: MAKYSSKINKIRTFALSLVFVGFLIMYIGVFFKESIWLSTFFMLLGVLSIGLSTVVYFWIGMLSTKAVRVVCPGCEKETKVLGRVDMCMHCREPLTLDPGLEGKEFDESYNRKKS . Based on its sequence composition, the protein contains hydrophobic regions suggesting possible membrane localization, and it includes several cysteine residues that may be involved in disulfide bond formation. Understanding the structural features of this protein is essential for determining its functional roles and potential applications in research.

How is recombinant BPUM_0828 typically produced in laboratory settings?

Recombinant BPUM_0828 protein is typically produced using an Escherichia coli expression system, particularly employing the BL21(DE3) strain or its derivatives, which are preferred hosts for recombinant protein production due to their fast growth, easy manipulation, and cost-effectiveness . The protein is commonly expressed with an N-terminal His-tag to facilitate purification through affinity chromatography techniques . The expression process generally involves cloning the BPUM_0828 gene into an expression vector under the control of a strong promoter such as the T7 promoter, which can be induced using isopropyl β-D-1-thiogalactopyranoside (IPTG) . The choice of expression system may need to be optimized considering factors such as codon usage, protein solubility, and potential toxicity to the host cell, with BL21(DE3) derivatives like C41(DE3) and C43(DE3) sometimes proving more suitable for proteins that may be challenging to express .

What are the recommended storage conditions for maintaining BPUM_0828 protein stability?

The optimal storage conditions for maintaining BPUM_0828 protein stability involve storing the lyophilized powder at -20°C or -80°C upon receipt, with aliquoting being necessary for multiple use scenarios to avoid repeated freeze-thaw cycles . For working solutions, it is advisable to store aliquots at 4°C for up to one week, as repeated freezing and thawing can lead to protein degradation and loss of activity . The protein is typically stored in a Tris/PBS-based buffer containing 6% trehalose at pH 8.0, which helps maintain stability during storage . When reconstituting the protein, it is recommended to briefly centrifuge the vial prior to opening to bring the contents to the bottom, and then reconstitute in deionized sterile water to a concentration of 0.1-1.0 mg/mL . Adding glycerol to a final concentration of 5-50% (with 50% being the default recommendation) before aliquoting can further enhance long-term storage stability at -20°C or -80°C .

What basic experimental controls should be included when working with recombinant BPUM_0828?

When designing experiments with recombinant BPUM_0828, several essential controls should be implemented to ensure reliability and validity of results. A negative control using a non-relevant protein with similar properties (size, tag system) should be included to distinguish specific interactions or effects from non-specific ones. Expression controls using samples collected before and after induction are crucial to confirm successful protein expression . Purification controls including analysis of flow-through, wash, and elution fractions help assess purification efficiency and protein quality. Buffer controls are necessary to evaluate the impact of buffer components on experimental outcomes, particularly important when the protein is stored in a buffer containing trehalose, which may affect certain assays . Activity controls utilizing proteins with known function related to the hypothesized function of BPUM_0828 provide a benchmark for assessing functional properties, while stability controls monitoring protein integrity over time under experimental conditions ensure that observed effects are not due to protein degradation.

What expression optimization strategies can overcome challenges in BPUM_0828 expression?

Optimizing BPUM_0828 expression may require a multi-faceted approach addressing several potential challenges. Codon optimization is a primary strategy, as differences in codon usage between Bacillus pumilus and E. coli can significantly impact expression efficiency; utilizing E. coli strains supplemented with rare tRNAs such as BL21(DE3) CodonPlus, Rosetta, or the integrated SixPack strain can help overcome codon bias issues . Expression conditions should be carefully optimized, including testing various induction temperatures (often lower temperatures of 16-25°C promote proper folding), IPTG concentrations, and induction durations to balance yield with proper folding . Alternative promoter systems may be beneficial if the T7 system leads to excessive expression or toxicity issues; the araBAD promoter offers more tunable expression compared to the powerful but less easily controlled T7 promoter . For membrane-associated proteins like BPUM_0828 (suggested by its sequence), specialized strains like C41(DE3) and C43(DE3) that can better tolerate membrane protein expression should be considered . If inclusion body formation occurs, co-expression with molecular chaperones or fusion to solubility-enhancing partners like thioredoxin or SUMO may improve soluble protein yield, while optimizing cell lysis and extraction conditions (detergents, solubilizing agents) can enhance recovery of membrane-associated proteins.

How can researchers design experiments to elucidate the function of the currently uncharacterized BPUM_0828 protein?

Designing experiments to elucidate the function of uncharacterized BPUM_0828 requires a comprehensive, multi-disciplinary approach starting with detailed bioinformatic analysis. Sequence-based predictions using tools like BLAST, Pfam, and structural prediction algorithms can identify conserved domains and potential functional motifs, while comparative genomics examining gene neighborhood and co-expression patterns across related species may reveal functional associations . Structural studies employing X-ray crystallography, NMR spectroscopy, or cryo-EM can provide insights into protein folding and potential binding sites, complemented by molecular dynamics simulations to predict dynamic properties and interaction potentials. Protein interaction studies using pull-down assays, yeast two-hybrid screens, or proximity labeling techniques can identify binding partners that may suggest functional roles. Knockout or knockdown studies in Bacillus pumilus coupled with phenotypic analysis can reveal physiological effects of protein absence, while heterologous expression followed by phenotypic characterization may identify gain-of-function effects. Functional reconstitution experiments with purified recombinant protein in defined biochemical assays testing various substrates and reaction conditions are essential for determining biochemical activity. Given BPUM_0828's membrane-associated characteristics and presence of cysteine residues, particular attention should be paid to potential roles in membrane processes, redox reactions, or metal binding, with appropriate assays designed to test these hypotheses .

What blocking strategies in experimental design are most effective when studying BPUM_0828 protein interactions?

When studying BPUM_0828 protein interactions, implementing effective blocking strategies in experimental design is crucial for obtaining reliable results by reducing variability and minimizing bias. Blocking by experimental batch is fundamental, ensuring that each treatment condition is represented in each experimental run to mitigate batch-to-batch variations in protein quality, reagent efficacy, or environmental conditions . Time-based blocking can help control for temporal fluctuations in laboratory conditions or equipment performance, particularly for experiments spanning multiple days or requiring time-sensitive measurements. Reagent-based blocking should be implemented when using different lots of antibodies, substrates, or other critical reagents by distributing these evenly across treatment groups . For experiments involving multiple protein preparations, blocking by preparation batch ensures that each treatment condition is tested with the same protein batches, controlling for variability in protein quality or activity between preparations. Technical skill-based blocking becomes important in multi-operator studies, distributing the work of different researchers evenly across treatment conditions to control for technique-based variability. In cell-based interaction studies, blocking by cell passage number or culture conditions helps control for variations in cellular response patterns. Properly implemented blocking strategies significantly enhance experimental power by reducing within-block variability, allowing for more precise detection of true interactions while utilizing fewer experimental units, thus saving both time and resources .

How can researchers address potential post-translational modification differences between native BPUM_0828 and the E. coli-expressed recombinant version?

Addressing post-translational modification (PTM) differences between native BPUM_0828 and its E. coli-expressed recombinant version requires both analytical approaches and potential expression system modifications. Initial comparative analysis using mass spectrometry-based proteomics should be performed on purified native protein from Bacillus pumilus and recombinant protein from E. coli to identify and characterize differences in PTMs such as phosphorylation, glycosylation, or disulfide bond formation . For critical disulfide bonds, the E. coli expression system can be modified by using strains with enhanced disulfide bond formation capabilities like Origami or SHuffle, or by directing the protein to the periplasm where the oxidizing environment favors disulfide formation . If phosphorylation is important, co-expression with relevant kinases from Bacillus pumilus or in vitro phosphorylation after purification may be necessary. For more complex modifications that E. coli cannot perform, alternative expression hosts such as Bacillus subtilis (closely related to B. pumilus), yeast systems, or mammalian cell lines might be more suitable, though these come with their own optimization challenges . Functional assays comparing the activity of native and recombinant proteins can help determine whether PTM differences significantly impact protein function. If maintaining specific PTMs is critical, chemical modification strategies or semi-synthetic approaches combining recombinant expression with chemical PTM addition might be considered. Researchers should always validate that the recombinant protein exhibits the expected biological activity despite potential PTM differences, and clearly document these considerations in experimental reports.

What quantitative methods are most appropriate for analyzing BPUM_0828 protein expression levels?

Quantitative analysis of BPUM_0828 protein expression levels can be approached through multiple complementary techniques, each with specific advantages. Western blotting with densitometry analysis provides a semi-quantitative assessment of protein expression using His-tag antibodies or specific antibodies against BPUM_0828, allowing comparison between different expression conditions while confirming the correct molecular weight and integrity of the expressed protein . More precise quantification can be achieved through ELISA-based methods, particularly suitable for comparing expression levels across multiple samples with high sensitivity, though this requires well-characterized antibodies against either the His-tag or BPUM_0828 itself. For absolute quantification, mass spectrometry-based approaches such as selected reaction monitoring (SRM) or parallel reaction monitoring (PRM) offer high specificity and sensitivity, enabling quantification even in complex mixtures by targeting specific peptides from BPUM_0828 . Fluorescence-based quantification methods such as using fluorescently labeled antibodies or expressing BPUM_0828 as a fusion with fluorescent proteins can enable real-time monitoring of expression levels in living cells, though care must be taken to ensure the fusion doesn't affect protein behavior. Spectrophotometric protein assays (Bradford, BCA, etc.) combined with SDS-PAGE analysis can provide total protein quantification along with an estimate of BPUM_0828 purity . For high-throughput screening of expression conditions, reporter systems linking BPUM_0828 expression to easily measurable outputs such as fluorescence or antibiotic resistance can be valuable, though they require validation with direct protein measurements .

What are the optimal E. coli strains and expression conditions for maximizing BPUM_0828 solubility?

Maximizing BPUM_0828 solubility requires careful selection of E. coli strains and expression conditions tailored to the protein's characteristics. Among E. coli strains, BL21(DE3) serves as a standard starting point due to its deficiency in lon and ompT proteases that can degrade recombinant proteins . For proteins with membrane-association potential like BPUM_0828, specialized strains such as C41(DE3) and C43(DE3) may offer advantages as they contain mutations that better accommodate membrane protein expression . If the protein contains multiple disulfide bonds (suggested by its cysteine content), strains such as Origami(DE3) or SHuffle with oxidizing cytoplasmic environments would be more suitable for proper folding . For optimal expression conditions, lower temperatures (16-25°C) often promote better folding by slowing down translation and allowing more time for proper folding, while reduced IPTG concentrations (0.1-0.5 mM rather than 1 mM) can prevent excessive expression that leads to aggregation . The composition of the growth medium significantly impacts solubility, with defined media or specialized formulations often yielding better results than standard LB media, particularly when supplemented with specific cofactors or metal ions that might be required for proper folding. Co-expression with molecular chaperones such as GroEL/ES, DnaK/J, or trigger factor can substantially increase soluble protein yields by assisting in the folding process . Addition of solubility-enhancing fusion partners such as MBP, SUMO, or thioredoxin can dramatically improve solubility, though these may need to be removed later depending on the experimental requirements . Induction at higher cell densities (OD600 of 0.6-0.8) often results in better soluble protein yields compared to induction at lower densities.

What purification strategy yields the highest purity and activity for recombinant His-tagged BPUM_0828?

A multi-step purification strategy is recommended to achieve highest purity and activity for recombinant His-tagged BPUM_0828. Initial capture should utilize immobilized metal affinity chromatography (IMAC) with Ni2+ or Co2+ resins, leveraging the N-terminal His-tag for selective binding; optimization of binding buffer composition (including salt concentration, pH, and potentially low concentrations of imidazole to reduce non-specific binding) is critical for this step . Following IMAC, size exclusion chromatography (SEC) serves as an excellent second purification step to separate the target protein from aggregates and impurities of different molecular sizes while simultaneously performing buffer exchange into a stabilizing formulation. Ion exchange chromatography (IEX) can be employed as an additional step if higher purity is required, with the choice between cation or anion exchange depending on the protein's isoelectric point relative to the working pH . For membrane-associated proteins like BPUM_0828, inclusion of appropriate detergents throughout the purification process is crucial for maintaining solubility and native conformation, with careful screening of detergent types and concentrations recommended. Throughout the purification process, activity assays should be performed on aliquots from each step to monitor retention of biological activity, guiding optimization of conditions to preserve functionality. The final purified product should undergo comprehensive quality assessment including SDS-PAGE, Western blotting, dynamic light scattering for aggregation analysis, and mass spectrometry for identity confirmation . For long-term storage, the purified protein should be formulated in a stabilizing buffer with appropriate additives such as trehalose (as mentioned in the product specifications) and stored as recommended at -20°C/-80°C with glycerol addition .

How should researchers approach troubleshooting low yield or insolubility issues with BPUM_0828 expression?

Addressing low yield or insolubility issues with BPUM_0828 expression requires a systematic troubleshooting approach starting with expression vector verification. Researchers should confirm the correct sequence through DNA sequencing and verify protein expression using small-scale test expressions with Western blotting to detect His-tagged products, checking for potential toxicity, premature termination, or degradation issues . Strain selection factors should be reassessed, potentially switching to specialized strains like C41(DE3)/C43(DE3) for potentially toxic proteins, or strains supplemented with rare tRNAs if codon usage analysis indicates potentially problematic rare codons in the BPUM_0828 sequence . Expression conditions should be systematically optimized through factorial design experiments varying temperature (typically testing 37°C, 30°C, 25°C, and 16°C), IPTG concentration (0.01-1 mM range), and induction time (2-24 hours), while also testing different media formulations including specialized media for membrane proteins if relevant . For persistent insolubility issues, solubility-enhancing strategies should be implemented, including fusion to solubility tags (MBP, SUMO, thioredoxin), co-expression with chaperones (GroEL/ES, DnaK/J systems), or directing expression to the periplasm if disulfide bonds are critical . If membrane association is suspected based on the protein sequence, appropriate detergents should be incorporated during extraction and purification, screening multiple detergent types (nonionic, zwitterionic) and concentrations . For proteins forming inclusion bodies, researchers might opt for optimized refolding protocols or consider native purification under denaturing conditions followed by controlled refolding if other approaches fail. Throughout this process, maintaining a detailed laboratory notebook recording all conditions tested and results obtained is essential for identifying patterns and determining the most promising approaches for optimization.

What statistical approaches are most appropriate for analyzing BPUM_0828 interaction data?

Analyzing BPUM_0828 interaction data requires robust statistical approaches appropriate for the specific interaction assay employed. For binding affinity measurements such as surface plasmon resonance (SPR) or isothermal titration calorimetry (ITC), nonlinear regression analysis using appropriate binding models (simple 1:1 binding, cooperative binding, or competitive binding) should be applied to determine key parameters like dissociation constants (Kd), with confidence intervals and goodness-of-fit metrics reported to assess reliability . Pull-down or co-immunoprecipitation assays should employ matched paired analysis with appropriate controls, using techniques like Student's t-test or ANOVA with post-hoc tests to compare binding between different conditions, while incorporating statistical corrections for multiple comparisons such as Bonferroni or false discovery rate methods . For large-scale interaction screenings like yeast two-hybrid or proximity labeling approaches, more sophisticated statistical treatments are necessary to distinguish true interactions from false positives, including significance analysis of interactome (SAINT) scoring, empirical determination of confidence score thresholds based on known controls, and enrichment analysis compared to appropriate negative controls . Experimental design should incorporate blocking strategies to minimize unwanted variation from batch effects, measurement time, or equipment differences, thereby improving statistical power to detect true interactions . Careful attention must be paid to assumptions underlying statistical tests, with data normality verified and non-parametric alternatives employed when necessary. Sample size determination should be performed a priori using power analysis based on expected effect sizes to ensure sufficient statistical power while avoiding excessive resource use. Visualization techniques including interaction networks, heat maps, and volcano plots can complement formal statistical analysis by revealing patterns that might not be apparent from numerical data alone.

How can researchers differentiate between specific and non-specific interactions when studying BPUM_0828?

Differentiating between specific and non-specific interactions in BPUM_0828 studies requires a multi-faceted validation approach with appropriate controls and characterization techniques. Comprehensive control experiments are fundamental, including using the His-tag alone (without BPUM_0828) as a negative control to identify interactions due solely to the tag, employing an unrelated protein of similar size and charge characteristics as another negative control, and including known interaction partners (if available) as positive controls . Dose-dependent binding analysis is crucial, as specific interactions typically show saturation kinetics with increasing protein concentration while non-specific interactions often increase linearly without saturation. Competition assays adding excess unlabeled protein to compete with labeled protein can confirm specificity, as true interactions will show decreased binding while non-specific interactions may not be affected. Mutational analysis targeting predicted interaction interfaces can validate binding mechanisms, as mutations disrupting key interface residues should significantly reduce specific interactions while having minimal effect on non-specific binding. Orthogonal binding assays employing different detection principles (e.g., SPR, ITC, FRET, co-IP) should be used to confirm interactions, as true interactions will typically be detected across multiple platforms while false positives may appear in only one. Stringency optimization through buffer composition adjustments (salt concentration, detergent type/concentration, pH) can help distinguish specific from non-specific interactions, with specific interactions often remaining stable under higher stringency conditions. For high-throughput studies, statistical approaches like significance analysis of interactome (SAINT) scoring or comparison to contaminant databases can help filter out common non-specific binders. Biological relevance assessment through techniques like co-localization studies, functional assays, or genetic interaction studies provides an additional layer of validation beyond biochemical interaction detection.

What approaches can resolve contradictory findings in BPUM_0828 functional characterization?

Resolving contradictory findings in BPUM_0828 functional characterization demands a methodical approach beginning with detailed method comparison. Researchers should analyze experimental protocols from conflicting studies, identifying differences in protein constructs, expression systems, purification methods, buffer conditions, and assay protocols that might explain divergent results . Rigorous replication efforts should be undertaken, reproducing the exact conditions from contradictory studies using the same reagents and protocols when possible, while implementing blind testing to minimize experimenter bias and establishing clear positive and negative controls to calibrate assay performance. Collaboration with groups reporting different findings enables direct comparison of materials and methods, potentially identifying subtle technical factors responsible for discrepancies. Expanding experimental approaches to include orthogonal techniques can provide independent verification, as truly robust findings should be detectable through multiple methodological approaches. Environmental and contextual factors should be systematically investigated, including testing protein function under varying conditions (pH, temperature, ionic strength, presence of cofactors) to determine if reported differences reflect condition-dependent activity rather than fundamental contradictions . Protein quality analysis using techniques like circular dichroism, thermal shift assays, or activity measurements on fresh versus stored samples can identify if stability or degradation issues contribute to contradictory findings. Meta-analysis techniques can be applied to aggregate data across multiple studies, potentially revealing patterns not apparent in individual reports. For complex contradictions, independent third-party validation by laboratories not previously involved in the conflicting studies offers an unbiased assessment. Throughout this process, researchers should maintain open communication with the scientific community through preprints, conference presentations, or direct exchanges to share insights on potential sources of variation and work collaboratively toward consensus.

How should researchers design control experiments when investigating potential binding partners of BPUM_0828?

Designing robust control experiments for BPUM_0828 binding partner investigations requires a comprehensive approach addressing multiple potential sources of false positives and negatives. Negative controls should include parallel experiments using the expression/purification tag alone (His-tag without BPUM_0828) to identify tag-mediated interactions, an unrelated protein of similar size, charge, and purification history to detect non-specific binding, and mock experiments with no bait protein to establish background binding levels to resins or matrices . Positive controls should be incorporated whenever possible, using known protein-protein interactions of similar strength and nature to validate assay functionality, though for novel proteins like BPUM_0828 with undefined function, engineered controls may need to be developed. Concentration-dependent controls are essential, testing a range of BPUM_0828 concentrations to distinguish specific interactions (which typically show saturation kinetics) from non-specific binding (which often increases linearly with concentration). Competitive binding experiments should be performed, where unlabeled BPUM_0828 competes with labeled protein for binding sites, with true interactions showing dose-dependent inhibition. Structure-based controls utilizing point mutations in predicted binding interfaces can provide powerful evidence for specific interactions, as mutations disrupting key interface residues should significantly reduce binding of true partners. Reciprocal experiments where the prey protein is used as bait and BPUM_0828 as prey provide additional validation, as true interactions should be detectable in both orientations. Stringency controls testing binding under increasing salt concentrations or detergent levels help distinguish robust interactions from weak or non-specific associations. Technical replications (using the same biological materials) establish measurement precision, while biological replications (independent protein preparations) confirm reproducibility across sample preparations . All controls should be performed under identical conditions to the test experiments, with samples processed in parallel and analyzed using the same methods and criteria.

What experimental design approach best elucidates the structure-function relationship of BPUM_0828?

Elucidating the structure-function relationship of BPUM_0828 requires an integrated experimental design combining structural analysis, functional assays, and mutational studies. The foundation should be high-resolution structural determination using X-ray crystallography, NMR spectroscopy, or cryo-electron microscopy to establish the three-dimensional architecture of the protein, complemented by computational modeling to predict functional sites based on structural features and evolutionary conservation . Based on structural information, a systematic site-directed mutagenesis campaign should target key residues selected through rational design principles, including conserved amino acids, predicted active sites, potential binding interfaces, and structural elements like the cysteine residues that may form disulfide bonds . The experimental design should follow a structure-guided hierarchical approach, first testing broader regional mutations (domain deletions or swaps) to identify critical functional regions, then focusing on specific residues within these regions to pinpoint essential amino acids. Parallel functional assays measuring multiple potential activities should be developed based on bioinformatic predictions and structural features, including potential enzymatic functions, protein-protein interactions, membrane association, and nucleic acid binding, with each assay optimized and validated using appropriate positive and negative controls . For each mutant, comprehensive characterization should include expression level, solubility, stability (using thermal shift assays), and structural integrity (using circular dichroism) to distinguish true functional defects from structural disruption. A factorial experimental design should be implemented to test mutants across varied conditions (pH, temperature, ionic strength, cofactor presence) to identify context-dependent functional effects that might reveal mechanistic insights . Complementation studies in relevant cellular systems (ideally B. pumilus or close relatives) can validate the biological relevance of identified structure-function relationships. Throughout this process, researchers should employ blocking strategies to control for batch effects and other sources of experimental variation, while ensuring sufficient biological and technical replication to support robust statistical analysis .

What future research directions are most promising for advancing understanding of BPUM_0828?

The advancement of understanding BPUM_0828 protein would benefit from several promising research directions that build upon current knowledge and leverage emerging technologies. High-resolution structural studies using X-ray crystallography, cryo-electron microscopy, or NMR spectroscopy would provide crucial insights into the protein's three-dimensional architecture, potentially revealing functional domains, binding sites, and structural features that could inform hypotheses about its biological role . Comprehensive interactome analysis employing techniques like proximity labeling (BioID, APEX), affinity purification-mass spectrometry, or protein microarrays could identify the protein's interaction partners in both Bacillus pumilus and heterologous systems, providing functional context through guilt-by-association . In vivo studies using knockout/knockdown approaches in Bacillus pumilus combined with phenotypic characterization would help establish the protein's physiological importance and potential cellular functions, complemented by localization studies using fluorescent protein fusions or immunofluorescence to determine subcellular distribution patterns. Evolutionary analysis examining the conservation, divergence, and co-evolution patterns of BPUM_0828 across bacterial species could provide insights into its functional importance and potential specialization. Systematic biochemical activity screening against diverse substrates, cofactors, and reaction conditions might identify enzymatic functions, while membrane interaction studies would be particularly relevant given the protein's hydrophobic regions suggesting potential membrane association . Integration of multiple omics approaches (transcriptomics, proteomics, metabolomics) examining changes associated with BPUM_0828 modulation could reveal affected pathways and processes. Development of specific antibodies against BPUM_0828 would facilitate numerous studies including detection, localization, and pull-down experiments. Computational approaches including molecular dynamics simulations and machine learning-based function prediction algorithms could generate testable hypotheses about the protein's behavior and interactions. Advanced genetic approaches such as suppressor screens or synthetic genetic array analysis might identify genes functionally related to BPUM_0828 through genetic interactions.

What are the key specifications of recombinant BPUM_0828 for research applications?

The key specifications of recombinant BPUM_0828 for research applications include detailed physical, chemical, and biological properties essential for experimental planning and execution. The complete table below summarizes these critical parameters:

This comprehensive specification profile provides researchers with the essential information needed for experimental planning, material handling, and results interpretation when working with recombinant BPUM_0828 protein . The detailed amino acid sequence allows for in silico analysis and primer design for further genetic manipulations, while storage and handling recommendations ensure maintenance of protein integrity throughout experimental workflows .