Recombinant Human Interleukin-17F (IL17F) (Active)

What is the structural composition of recombinant human IL-17F?

Recombinant human IL-17F is a full-length protein spanning amino acids 31-163, with a complete size of 163 amino acids including a 30 amino acid signal peptide. The mature protein forms disulfide-linked dimers and shares approximately 44% amino acid sequence homology with IL-17A, while having only limited sequence homology (16-30%) with other IL-17 family members (IL-17B, C, D and E). Human and mouse IL-17F share 55% sequence identity . The protein contains a conserved cystine-knot fold near the C-terminus, which is characteristic of the IL-17 family .

How does IL-17F differ from other IL-17 family members in terms of receptor binding?

IL-17F primarily signals through the IL-17RA-IL17RC heterodimeric receptor complex, though IL-17F homodimers can also signal via IL17RC homodimeric receptor complexes . While IL-17 RA binds IL-17A and IL-17 RB binds IL-17B and IL-17E, IL-17 RC binds both IL-17A and IL-17F with similarly high affinity, functioning as a receptor for both cytokines . This binding triggers homotypic interaction of IL17RA and IL17RC chains with TRAF3IP2 adapter through SEFIR domains, leading to downstream signal transduction . This receptor binding profile distinguishes IL-17F's biological activity from other family members.

What are the primary cellular sources of IL-17F in the human body?

IL-17F is predominantly expressed in activated CD4+ T cells (particularly T-helper 17 cells) and activated monocytes . As a signature effector cytokine of T-helper 17 cells, IL-17F primarily induces neutrophil activation and recruitment at infection and inflammatory sites . Additionally, other innate immune cells including neutrophils, monocytes, and macrophages can produce IL-17F in specific inflammatory contexts .

What is the canonical signaling pathway activated by IL-17F?

IL-17F signaling is initiated when it binds to the IL-17RA-IL17RC heterodimeric receptor complex. This binding triggers the recruitment of the adapter protein Act-1 (TRAF3IP2) through SEFIR domain interactions, followed by TRAF6 recruitment. The signaling cascade continues with downstream activation of NF-κB and MAPK pathways . Recent research has shown that nIL-17 (the bioactive 20-mer sequence of IL-17) amplifies the expression of both Act-1 and NFκB without altering IL-17RA or IL-17RC receptor expression levels . This ultimately results in the transcriptional activation of cytokines, chemokines, antimicrobial peptides, and matrix metalloproteinases, potentially triggering strong inflammatory responses .

What is the newly identified bioactive sequence (nIL-17) and how does it relate to IL-17F function?

Researchers have recently identified a critical 20-mer amino acid sequence, named nIL-17, which is responsible for the biological activity of both IL-17A and IL-17F. This peptide effectively mimics the C-terminal region of IL-17A/F that is essential for receptor interaction. In experimental settings, nIL-17 has demonstrated the ability to induce IL-6 production in NIH-3T3 mouse embryonic fibroblast cells to a similar or greater extent than both full-length IL-17A and IL-17F at comparable molar concentrations . nIL-17 binds to both IL-17RA and IL-17RC receptors with similar efficacy as the native proteins, activating downstream signaling pathways and inducing inflammatory responses comparable to full-length IL-17A/F .

How does IL-17F contribute to antimicrobial host defense?

IL-17F plays a crucial role in antimicrobial host defense through multiple mechanisms. It regulates the composition of intestinal microbiota and immune tolerance by inducing antimicrobial proteins that specifically control the growth of commensal Firmicutes and Bacteroidetes . IL-17F stimulates mucosal epithelial cells to produce antimicrobial beta-defensins (DEFB1, DEFB103A, and DEFB104A), which limit microbe entry through epithelial barriers . Additionally, IL-17F is primarily involved in defense against extracellular bacteria and fungi by inducing neutrophilic inflammation, activating neutrophils, and promoting their recruitment to infection sites . These mechanisms collectively contribute to the cytokine's essential role in protecting host tissues from microbial invasion.

What are the optimal experimental conditions for studying IL-17F-induced cellular responses?

When studying IL-17F-induced cellular responses, researchers should consider several key experimental parameters. For in vitro studies, NIH-3T3 mouse embryonic fibroblasts and human dermal blood endothelial cells (HDBECs) have been successfully used to study IL-17F signaling . Effective concentrations of recombinant IL-17F range from 0.610 nM to 0.725 nM for inducing measurable cellular responses . When assessing inflammatory amplification, M1 macrophages rather than M0 or M2 macrophages should be used, as IL-17F specifically increases IL-6 and TNF-α release from the M1 phenotype . For receptor binding assays, biotinylated IL-17F can be used to assess interactions with both IL-17RA and IL-17RC . Cell-based assays should include appropriate controls to distinguish IL-17F-specific effects from background signaling.

How can researchers accurately measure IL-17F-induced signaling pathway activation?

To accurately measure IL-17F-induced signaling pathway activation, researchers can employ multiple complementary approaches. Western blotting can be used to detect phosphorylation of downstream signaling molecules in the NF-κB and MAPK pathways following IL-17F stimulation . Quantitative PCR is effective for measuring changes in gene expression of Act-1 and NF-κB components, as well as target genes induced by IL-17F signaling . ELISA assays can quantify the production of inflammatory mediators such as IL-6, IL-8, TNF-α, and G-CSF in response to IL-17F stimulation . For cellular responses, researchers can assess neutrophil migration, adhesion molecule expression on endothelial cells, and fibroblast activation . Comparing responses between full-length IL-17F and the nIL-17 peptide can provide additional insights into signaling mechanisms .

What experimental models are most appropriate for studying IL-17F functions in inflammatory diseases?

For studying IL-17F functions in inflammatory diseases, several experimental models have proven valuable. In vitro, NIH-3T3 fibroblasts and human M1 macrophages can be used to study IL-17F-mediated inflammatory amplification . For in vivo studies, the air pouch model effectively demonstrates IL-17F's role in leukocyte recruitment to pre-inflamed tissues . Mouse models of rheumatoid arthritis and inflammatory bowel disease have been successfully employed to evaluate the therapeutic efficacy of antibodies targeting IL-17F signaling . When selecting a model, researchers should consider that IL-17F functions may vary by tissue context—effects in dermal, joint, and intestinal tissues might differ significantly . Additionally, comparing IL-17F-deficient and wild-type animals in disease models can elucidate the specific contributions of this cytokine to pathophysiology.

How does IL-17F cooperate with other cytokines in inflammatory disease pathogenesis?

IL-17F exhibits complex cooperative relationships with multiple cytokines in inflammatory disease contexts. While IL-17F alone can induce certain inflammatory responses, its effects are significantly amplified when combined with TNF-α, IL-6, or IL-1β . IL-17F works synergistically with TNF-α to enhance the production of inflammatory mediators from fibroblasts and endothelial cells beyond what either cytokine induces individually . In macrophages, IL-17F specifically amplifies cytokine production from M1 (but not M0 or M2) macrophages, suggesting context-dependent cooperative effects . The IL-17F homodimer exhibits functional overlap with IL-17A homodimers and IL-17A/F heterodimers, though generally with lower potency than IL-17A . Understanding these cooperative interactions is crucial when designing experimental approaches to study inflammatory disease mechanisms and when developing therapeutic strategies targeting IL-17-mediated inflammation.

What are the key differences between IL-17F and IL-17A in experimental systems?

Despite their structural similarities, IL-17F and IL-17A exhibit important functional differences in experimental systems. IL-17F shares approximately 44% amino acid sequence homology with IL-17A but generally demonstrates lower inflammatory potency . While IL-17A functions as a direct chemotactic agent for neutrophils, IL-17F exhibits this property to a lesser extent . Both cytokines signal through the IL-17RA/RC receptor complex, but IL-17F can also signal via IL-17RC homodimers . IL-17A and IL-17F induce overlapping but distinct gene expression profiles in target cells, with IL-17A typically inducing stronger inflammatory responses . The recently identified bioactive nIL-17 peptide mimics the activities of both cytokines but shows comparable binding profiles to both receptors . These differences highlight the importance of cytokine-specific experimental approaches when studying IL-17 family members.

How can the nIL-17 peptide be utilized in advanced IL-17F research applications?

The recently identified nIL-17 bioactive peptide offers several innovative applications for advanced IL-17F research. As a mimetic of the active site of both IL-17A and IL-17F, nIL-17 can be used as a tool to study receptor binding and activation mechanisms without the complexities of full-length protein dimers . Researchers can employ structure-activity relationship studies with modified versions of nIL-17 (such as nIL-17A-NH₂, nIL-17A-DN, or nIL-17A-SC) to identify critical amino acid residues required for receptor interaction and signaling . The peptide can serve as a standardized reagent for comparing IL-17A and IL-17F activities across experimental systems. Additionally, nIL-17 has potential as a screening tool for identifying novel IL-17 pathway inhibitors in drug discovery platforms . The development of neutralizing antibodies against nIL-17 (such as Ab-IPL-IL-17) presents opportunities for more targeted therapeutic approaches with potentially reduced immunogenicity compared to antibodies against full-length proteins .

How do neutralizing antibodies against IL-17F differ from those targeting other IL-17 family members?

Neutralizing antibodies against IL-17F exhibit distinct characteristics compared to those targeting other IL-17 family members. Unlike secukinumab and ixekizumab, which specifically target IL-17A, antibodies against IL-17F must contend with its unique structural features while sharing the 44% sequence homology with IL-17A . Bimekizumab targets both IL-17A and IL-17F, recognizing shared epitopes . The recently developed Ab-IPL-IL-17, which targets the bioactive nIL-17 peptide sequence found in both IL-17A and IL-17F, shows promising neutralizing activity against both cytokines . Importantly, studies indicate that antibodies targeting the nIL-17 sequence may exhibit lower immunogenicity and fewer adverse hematological side effects compared to reference antibodies against full-length proteins . This suggests that targeting the bioactive peptide region rather than the complete protein structure may offer therapeutic advantages in inflammatory disease treatment.

What role does IL-17F play in different immune-mediated inflammatory diseases (IMIDs)?

IL-17F contributes distinctively to various immune-mediated inflammatory diseases (IMIDs). In rheumatoid arthritis, IL-17F promotes inflammation in the synovium by stimulating fibroblast-like synoviocytes to produce inflammatory mediators and matrix-degrading enzymes . In psoriasis, IL-17F acts on keratinocytes, contributing to the characteristic epidermal hyperplasia and inflammatory infiltrate . For inflammatory bowel disease, IL-17F plays a dual role—while it helps maintain epithelial barrier function by inducing antimicrobial peptides, dysregulated IL-17F signaling can amplify pathological inflammation . In all these conditions, IL-17F works cooperatively with other cytokines, particularly TNF-α, to enhance inflammatory responses . Unlike IL-17A, which shows stronger associations with disease severity, IL-17F often plays more subtle, context-dependent roles in IMIDs . Current therapeutic approaches targeting both IL-17A and IL-17F (like bimekizumab) have shown efficacy in treating plaque psoriasis, psoriatic arthritis, and ankylosing spondylitis .

How can researchers design experiments to evaluate IL-17F-targeted therapeutics?

When designing experiments to evaluate IL-17F-targeted therapeutics, researchers should implement a comprehensive approach across multiple experimental systems. In vitro screening should begin with binding assays to confirm target engagement, using both recombinant IL-17F and the nIL-17 bioactive peptide to assess binding specificity and affinity . Functional assays using NIH-3T3 fibroblasts or HDBECs can evaluate the therapeutic's ability to neutralize IL-17F-induced IL-6, IL-8, or adhesion molecule expression . For more complex cellular systems, researchers should assess the therapeutic's effects on IL-17F-mediated inflammatory amplification in primary human M1 macrophages . In vivo evaluation should employ relevant disease models, such as arthritis or inflammatory bowel disease models, comparing the candidate therapeutic to reference anti-IL-17 antibodies . Importantly, researchers must monitor both efficacy (reduction in inflammatory markers and disease progression) and safety (immunogenicity and hematological parameters) to fully characterize the therapeutic profile . Combination studies with other anti-cytokine therapeutics can provide insights into potential synergistic treatment strategies.

What are the critical quality attributes to verify when using recombinant IL-17F in research?

When using recombinant IL-17F in research, several critical quality attributes must be verified to ensure experimental reliability. Researchers should confirm protein purity (≥95% is recommended) through SDS-PAGE or other appropriate analytical methods . Endotoxin levels should be assessed and confirmed to be ≤0.005 EU/μg to prevent confounding inflammatory responses in experimental systems . Biological activity verification is essential and can be performed by measuring IL-6 induction in NIH-3T3 cells or similar bioassays . The dimeric state of the protein should be confirmed, as IL-17F functions as a disulfide-linked dimer . For expression system considerations, recombinant IL-17F produced in HEK293 cells provides appropriate post-translational modifications for human research applications . Researchers should verify lot-to-lot consistency when conducting longitudinal studies to minimize experimental variability.

How should researchers troubleshoot inconsistent results in IL-17F signaling experiments?

When encountering inconsistent results in IL-17F signaling experiments, researchers should systematically evaluate several key factors. First, verify recombinant IL-17F quality through fresh aliquot preparation, as repeated freeze-thaw cycles can compromise protein activity . Check receptor expression levels in the experimental cell system, as IL-17RA and IL-17RC expression can vary between cell types and culture conditions . For signaling pathway analysis, include positive controls (such as TNF-α) to confirm cell responsiveness, and consider that IL-17F generally induces weaker responses than IL-17A in many systems . If using primary cells, donor-to-donor variability may significantly impact results, necessitating increased biological replicates . The cellular activation state is crucial, particularly for macrophages, where IL-17F specifically amplifies responses in M1 (but not M0 or M2) macrophages . Finally, consider potential synergistic effects with other cytokines present in the experimental system, as IL-17F often works cooperatively with TNF-α and other inflammatory mediators .

What are the optimal storage and handling conditions for maintaining IL-17F biological activity?

To maintain optimal IL-17F biological activity, researchers should adhere to specific storage and handling protocols. Recombinant IL-17F should be stored at -80°C for long-term preservation and at -20°C for shorter periods . The protein should be aliquoted upon receipt to minimize freeze-thaw cycles, as repeated freezing and thawing significantly reduces biological activity . When preparing working solutions, IL-17F should be diluted in appropriate buffers containing carrier protein (such as 0.1% BSA) to prevent adhesion to container surfaces and maintain stability . During experiments, IL-17F solutions should be kept on ice and used within the same day of preparation . For maximum stability, avoid exposing the protein to extreme pH conditions, oxidizing agents, or proteases . When performing long-term studies, researchers should consider conducting parallel bioactivity assays using reference standards to confirm consistent protein activity throughout the experimental timeline.