Detection of Human PACT by Western Blot. Western blot shows lysates of K562 human chronic myelogenous leukemia cell line and HeLa human cervical epithelial carcinoma cell line. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human PACT Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1980) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for PACT at approximately 32 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Detection of Human PACT by Simple WesternTM. Simple Western lane view shows lysates of K562 human chronic myelogenous leukemia cell line, loaded at 0.2 mg/mL. A specific band was detected for PACT at approximately 42 kDa (as indicated) using 10 µg/mL of Goat Anti-Human PACT Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1980) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.
Preparation and Storage
Reconstitute at 0.2 mg/mL in sterile PBS.
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
PACT (PKR Activator; also HSD14, PRKRA and
RAX [in mouse]) is a 34-37 kDa member of the PRKRA family of molecules. It is a ubiquitously expressed, cytoplasmic,
313 amino acid (aa) protein that contains three discrete double-stranded (ds)
RNA binding motifs (aa 31-101; 126-194; 240-308). It exists as a monomer, homodimer, and
heterodimer with at least two partners, TRBP and Dicer. Two isoform variants are known to exist,
including one that utilizes an alternative start site at Met26, and another
that shows a 10 aa substitution for aa 1-21.
PACT is perhaps best known for its relationship with PKR, a 52-58 kDa Ser/Thr
kinase that downmodulates protein synthesis and initiates apoptosis. PKR is constitutively silent, and activated
via two principal ways; the first is through binding to (viral) dsRNA, and the
second is through binding to an activator, PACT. PACT is kept in check upstream through
dimerization with at 48-49 kDa regulatory protein termed TRBP. Following growth factor withdrawl or exposure
to bacterial products such as LPS, PACT is phosphorylated at Ser287, and this
inactive PACT:TRBP heterodimer dissociates, freeing up PACT to bind to
PKR. PACT's interaction with PKR induces
an autophosphorylation event on PKR.
This allows for an activated PKR interaction with eIF2a, causing
its phosphorylation and a subsequent reduction of protein synthesis. PACT may also act independently of PKR, and
serve as a component of RISC, a complex that downregulates mRNA
translation. Within this complex, PACT
likely associates with Dicer in concert with Argonaute. Full-length PACT shares 98% aa sequence
identity with the mouse ortholog to human PACT termed RAX.
Entrez Gene IDs:
8575 (Human); 23992 (Mouse); 311130 (Rat)
DYT16; DYT16interferon-inducible double stranded RNA-dependent activator; HSD14; PACTPKR-associating protein X; PKR-associated protein X; PKR-Associating Protein X; PRKRA; protein kinase, interferon-inducible double stranded RNA dependent activator; RAX; RAXHSD14