KLRK1 Human, Sf9

What is KLRK1 and what is its role in immune surveillance?

KLRK1/NKG2D is an activating receptor primarily expressed on Natural Killer (NK) cells and certain T cell subsets. It plays a crucial role in immune surveillance by recognizing stress-induced ligands like MICA and MICB that are expressed during cell stress and up-regulated in tumor cells and viral infections . The receptor-ligand interaction between KLRK1 and these stress proteins has essential functions in immune responses to various pathologies, including cancer and viral infections .

In experimental settings, KLRK1 Human produced in Sf9 baculovirus cells allows researchers to study receptor-ligand interactions and downstream signaling pathways. The recombinant protein contains 386 amino acids (73-216 a.a.) with a molecular mass of 43.9kDa and typically appears at approximately 40-57kDa on SDS-PAGE .

What signaling pathways are activated by KLRK1 in different immune cell populations?

KLRK1 signaling varies between different immune cell populations:

• In NK cells: KLRK1 engagement leads to activation of cytotoxic functions and cytokine production, particularly through interaction with adaptor molecules

• In T cells: KLRK1 acts as a co-stimulatory receptor that enhances TCR-mediated responses, with significant upregulation of activation-related genes including Cd69, Cd226, Cd40lg and Klrk1 itself

Research using KLRK1 Human, Sf9 has helped elucidate that CD4+ T cells with activated KLRK1 signaling show a strong T helper type 1 signature with significantly higher expression of Tbx21 and Ifng, important for anti-tumor responses .

How do KLRK1 and its ligands contribute to anti-tumor immunity?

KLRK1/NKG2D makes significant contributions to anti-tumor immunity through several mechanisms:

• Recognition of stress-induced MICA/MICB proteins that are upregulated on tumor cells

• Activation of both NK cells and specific T cell subsets within the tumor microenvironment

• Recruitment of multiple immune effector populations to tumors

Studies have shown that targeting the MICA/B-KLRK1 axis through vaccination approaches can result in significant tumor infiltration by immune cells - up to 17.9-fold enrichment in CD8+ T cells and a 38.9-fold enrichment in NK cells compared to control vaccinations . This dual activation of both adaptive (T cell) and innate (NK cell) immune compartments makes KLRK1 particularly valuable for cancer immunotherapy research.

What are the optimal storage and handling conditions for KLRK1 Human, Sf9?

For optimal preservation of KLRK1 Human, Sf9 activity:

• Storage solution: Keep the protein in Phosphate Buffered Saline (pH 7.4) with 10% glycerol

• Long-term storage: For extended periods, add a carrier protein (0.1% HSA or BSA) to prevent degradation

• Temperature sensitivity: Avoid multiple freeze-thaw cycles which can compromise protein integrity

• Purity considerations: Work with preparations that show >95.0% purity as determined by SDS-PAGE

These conditions ensure the protein maintains its native conformation and binding capacity for experimental applications.

How can researchers validate the functional activity of KLRK1 Human, Sf9?

Validation of functional KLRK1 Human, Sf9 should incorporate:

• Binding assays with known ligands (MICA/MICB)

• Comparison to native receptor activity in primary NK cells

• Verification that the C-terminal tag (242 amino acids hIgG-His tag) does not interfere with binding functionality

• Flow cytometry to confirm interaction with ligand-expressing cells

• Functional assays measuring activation of KLRK1-expressing immune cells

Researchers should recognize that the recombinant protein contains the amino acid sequence ADPIWSAVFLNSLFNQEVQIPLTESYCGPCPKNWICYKNNCYQFFDESKNWYESQASCMSQNASLLKVYSKEDQDLLKLVKSYHWMGLVHIPTNGSWQWEDGSILSPNLLTIIEMQKGDCALYASSFKGYIENCSTPNTYICMQRTVVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV , which includes both the receptor domain and the tag structure.

What considerations are important when using KLRK1 Human, Sf9 in cell-based assays?

When incorporating KLRK1 Human, Sf9 into cell-based experimental systems:

• Buffer composition: Use physiologically relevant buffers that maintain protein stability

• Concentration titration: Establish dose-response relationships (typically 0.1-10μg/ml range)

• Controls: Include heat-inactivated protein and blocking antibodies to confirm specificity

• Timing: Assess both early activation markers (4-6 hours) and later functional responses (12-24 hours)

• Tag considerations: Be aware that the hIgG-His tag may interact with Fc receptors on target cells

These methodological details are critical for generating reproducible and physiologically relevant results when studying KLRK1-mediated cellular responses.

How is KLRK1 Human, Sf9 being used in cancer vaccine development?

KLRK1 Human, Sf9 has become an important tool in developing cancer vaccines that target tumor immune escape mechanisms. Recent research has created vaccines targeting MICA and MICB stress proteins that are recognized by KLRK1/NKG2D . These approaches have demonstrated:

• Significant enrichment of tumor-infiltrating T cells and NK cells

• Reduction in FoxP3+ regulatory T cells within tumors

• Higher NKG2D and CXCR6 receptor levels on tumor-infiltrating CD4+ and CD8+ T cells

• Substantial clonal expansion of both CD4+ and CD8+ T cell populations

• Activation of three clusters of group 1 innate lymphoid cells (ILCs): ILC1s, Xcl1+ NK cells and cytotoxic NK cells

The experimental use of KLRK1 Human, Sf9 helps in validating these vaccine constructs through binding studies and functional characterization.

What role does KLRK1 play in the cellular innate immune response?

KLRK1 represents an important component of the cellular innate immune response, which is activated in response to various threats. This system includes:

• Recognition of foreign materials like synthetic RNAs

• Expression of Type I or Type II interferons

• Induction of IFN signature genes including IFNα, IFNB1, IFIT, OAS1, PKR, RIGI, and others

Understanding how KLRK1 integrates into this broader immune surveillance network is essential for researchers developing strategies to either enhance immune responses (for cancer) or modulate excessive immune activation (for autoimmune conditions).

How does KLRK1 function differ between NK cells and T cell subsets?

The functional activity of KLRK1 shows important differences between immune cell populations:

Research shows that CD4+ and CD8+ T cells utilize KLRK1/NKG2D somewhat differently than NK cells, with T cells showing higher expression of chemokine receptors like CXCR3 and CXCR6 following activation . This allows precise trafficking to inflammatory sites where KLRK1 ligands may be expressed.

How can researchers overcome challenges with protein stability?

When encountering stability issues with KLRK1 Human, Sf9:

• Add carrier proteins (0.1% HSA or BSA) to prevent adsorption to container surfaces

• Prepare small aliquots to avoid repeated freeze-thaw cycles

• Use low-binding microcentrifuge tubes and pipette tips to minimize protein loss

• Consider adding additional stabilizers if extended experimental protocols are necessary

• Monitor protein integrity via SDS-PAGE before critical experiments

The formulation described in the technical information (Phosphate Buffered Saline with 10% glycerol) provides a starting point, but may require optimization for specific applications .

What approaches help distinguish between specific and non-specific effects when using KLRK1 Human, Sf9?

To ensure experimental results reflect genuine KLRK1-mediated effects:

• Include blocking antibodies specific to the KLRK1 receptor domain

• Test graduated concentrations to establish dose-dependence

• Use control proteins with similar tag structures but different receptor domains

• Compare results with native KLRK1 expression systems when possible

• Consider the potential impact of the C-terminal tag on protein function

These controls are particularly important when studying novel KLRK1 interactions or when developing new experimental systems.

How should researchers interpret binding data between KLRK1 and various stress-induced ligands?

When analyzing binding interactions between KLRK1 Human, Sf9 and stress ligands:

• Consider the hierarchical binding patterns typically observed (MICA/B often showing higher affinity than ULBPs)

• Account for allelic variations in ligands that can substantially affect binding affinity

• Recognize that insect cell-derived KLRK1 may have different glycosylation patterns than mammalian-expressed protein

• Validate key findings using cell-based systems with native receptor expression

• Report comparative binding parameters rather than absolute values when using the recombinant system

How might KLRK1 research contribute to understanding autoimmune and inflammatory diseases?

While KLRK1 research has centered largely on cancer immunity, emerging evidence suggests important roles in autoimmune pathology:

• Potential involvement in conditions like Behçet's Disease, where genetic susceptibility may involve immune receptors under evolutionary selection pressure

• Possible connection to dementia pathophysiology, where immune receptor genes show differential expression patterns

• Contribution to inflammatory cascades that have both protective and pathological potential

Future research using KLRK1 Human, Sf9 may help characterize how variations in this receptor contribute to disease susceptibility and progression across various inflammatory conditions.

What innovative technologies are advancing KLRK1 functional studies?

Cutting-edge methodologies enhancing KLRK1 research include:

• Single-cell transcriptomics revealing receptor-associated gene networks

• CRISPR-Cas9 gene editing to precisely manipulate KLRK1 expression or function

• Advanced imaging techniques visualizing KLRK1 clustering and signaling dynamics

• Proteomics approaches identifying novel KLRK1-associated proteins

• Structural biology determining binding interfaces at atomic resolution

The controlled nature of recombinant KLRK1 Human, Sf9 makes it particularly valuable for standardizing these advanced technological applications.

How might synthetic RNA technologies interface with KLRK1 research?

Emerging RNA technologies present interesting opportunities for KLRK1 research:

• Modified synthetic RNAs that avoid triggering cellular innate immune responses

• RNA-based therapeutics potentially targeting KLRK1 expression or function

• RNA vaccines encoding KLRK1 ligands for cancer immunotherapy

These approaches could leverage the modifications described in immunotherapy research, including 5-methylcytidine, N6-methyladenosine, 2-thiouridine, and other nucleoside modifications that avoid triggering interferon responses .