PKR Kinase Activation Assay Kit

The PKR Kinase Activation Assay Kit is designed to measure the activation of human erythrocyte pyruvate kinase enzyme PKR, also known as PKLR Var 1, for screening and profiling applications using Kinase-Glo(R) Max as a detection reagent. The assay kit comes in a convenient 96-well format, with enough purified PKR, Phosphoenolpyruvate (PEP), ADP, allosteric PK activator FBP (fructose-1,6-bisphosphate), and kinase assay buffer for 100 enzyme reactions. Need us to run activator screens or profile your activators against PKR? Check out our Kinase Screening Services. Red blood cells rely entirely on glycolysis for ATP production, making the erythroid pyruvate kinase isoform PKR (PKLR Var1) essential for maintaining cellular energy and integrity, reduced PKR activity lowers ATP and causes accumulation of glycolytic intermediates such as 2,3 diphosphoglycerate (2,3 DPG), which increases hemoglobin S (HbS) polymerization, promotes sickling, and contributes to chronic hemolytic anemia. PK deficiency-the most common cause of nonspherocytic hemolytic anemia-shares this metabolic signature of elevated 2,3 DPG and reduced ATP, similar to sickle RBCs, contributing to anemia, dehydration, and increased sickling propensity. With more than 290 PKLR mutations identified, several linked to acute pain in sickle cell disease (SCD), these metabolic disturbances provide a strong rationale for targeting PKR. The tetrameric R form of red cell PK (PKR) has naturally low affinity for PEP, which is increased upon binding of fructose 1,6 bisphosphate (FBP), its primary allosteric activator, FBP increases PEP affinity, promotes tetramerization, and stabilizes PKR in its active form. Recently developed oral allosteric activators-mitapivat and etavopivat-bind a pocket at the dimer-dimer interface distinct from the FBP site and increase PKR activity in both wild type and mutant enzymes. Etavopivat and mitapivat have been shown to decrease 2,3 DPG, increase ATP, raise hemoglobin oxygen affinity, and reduce sickling in sickle RBCs, confirming the therapeutic potential of PKR activation.