Pam3CSK4

Synthetic triacylated lipopeptide

TLR2/TLR1 activation with Pam3CSK4

Pam3CSK4 (Pam3CysSerLys4) is a synthetic triacylated lipopeptide (LP) and a TLR2/TLR1 ligand. It is a potent activator of the pro-inflammatory transcription factor NF-κB [1, 2]. Pam3CSK4 mimics the acylated amino terminus of bacterial LPs.

Bacterial LPs are a family of pro-inflammatory cell wall components found in both Gram-positive and Gram-negative bacteria. The stimulatory activity of these LPs resides in their acylated amino terminus. These bacterial LPs are recognized by TLR2, a receptor that plays a pivotal role in detecting a diverse range of pathogen-associated molecular patterns (PAMPs) [3].

At the cell surface, TLR2 forms a heterodimer with co-receptors TLR1 or TLR6, depending upon either tri- or diacylation of the ligand. Once a ligand binds to either TLR2-TLR1 or TLR2-TLR6, a MyD88-dependent activation of NF-κB and AP-1 occurs, ultimately leading to an innate immune response. Recognition of Pam3CSK4, a triacylated LP, is mediated by TLR2 which cooperates with TLR1 through their cytoplasmic domain to induce the signaling cascade leading to the activation of NF-κB [4].

Key Features of Pam3CSK4:

• Potent activator of the TLR2/TLR1 heterodimer
• Synthetic lipopeptide free of microbial contaminants
• Each lot is highly pure (≥95%) and functionally tested

Read our review on TLR2.

References:

1. Brandt K.J. et al., 2013. TLR2 Ligands Induce NF-κB activation from endosomal compartments of human monocytes. PLoS One. 8(12) :e80743.
2. Aliprantis A.O. et al., 1999. Cell activation and apoptosis by bacterial lipoproteins through toll-like receptor-2. Science 285(5428) :736-9.
3. Oliveira-Nascimento L. et al., 2012. The Role of TLR2 in Infection and Immunity. Front Immunol. 3 :79.
4. Ozinsky A. et al.., 2000. The repertoire for pattern recognition of pathogens by the innate immune system is defined by cooperation between toll-like receptors. PNAS. 97(25) :13766-71.

Figures

Pam3CSK4 induces a dose-dependent response in HEK-Blue™ hTLR2 cells. These cells were stimulated with increasing concentrations of Pam3CSK4. After overnight incubation, the NF-κB response was determined using HEK-Blue™ Detection, a SEAP detection medium, and by reading the optical density (OD) at 630 nm. 

Specific TLR2-mediated NF-κB activation by Pam3CSK4. THP1-Dual™ and THP1-Dual™ KO-TLR2 cells were stimulated with increasing concentrations of Pam3CSK4. After overnight incubation, the NF-κB response was determined using QUANTI-Blue™ Solution, a SEAP detection reagent, and by reading the optical density (OD) at 630 nm. The IRF response was assessed by measuring the activity of Lucia luciferase in the supernatant using QUANTI-Luc™. Data for the Lucia luciferase readout are shown as a fold increase over non-induced cells. As expected, THP1-Dual™ KO TLR2 cells did not respond to Pam3CSK4. Of note, due to TLR2 not directly signaling through an IRF-dependent pathway, Pam3CSK4  did not induce a significant fold-increase in the IRF-inducible reporter Lucia luciferase in the THP1-Dual™ cells.

Specifications

Specificity: TLR2/TLR1 agonist

Working concentration: 0.1 - 10 ng/ml

CAS number: 112208-01-2

Molecular formula: C₈₁H₁₅₆N₁₀O₁₃S • 3TFA

Molecular weight: 1852.33 g/mol

Solubility: 2 mg/ml in water

Quality control:

• Purity: ≥95% (UHPLC)
• Activation of TLR2 by Pam3CSK4 has been confirmed using cellular assays.
• The absence of endotoxins has been confirmed using HEK-Blue™ hTLR4 cells.

Contents

Pam3CSK4 is provided lyophilized.

• 1 mg of Pam3CSK4
• 1.5 ml of endotoxin-free water

Pam3CSK4 is shipped at room temperature.

Upon receipt, store at 4°C.

Lyophilized Pam3CSK4 is stable for 1 year when properly stored.