Myc/his-tagged WT or S208A/S242A mutant N-terminal pyrin (aa’s 1- 330) was overexpressed into 293T cells, and the N-terminal pyrin proteins were purified using His-SpinTrap™ (28-4013-53, GE Healthcare). Each purified N-terminal pyrin protein was incubated with 5 μg of PKN1 (PR7255B) or PKN2 (PR7370A, Thermo Scientific) at 30°C for 0.5h in a kinase buffer (50 mM HEPES pH7.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA, 500 μM ATP and 2.5 mM DTT). The reactants were analyzed by immunoblotting with anti-phospho-Ser antibody (9606, Cell Signaling) or staining with Pro-Q diamond (P33300, Invitrogen). Pro-Q diamond-stained gel was visualized on Molecular Imager FX (Bio-Rad).
Anti phospho ser antibody
Manufactured by Cell Signaling Technology
The Anti-phospho-Ser antibody is a laboratory reagent designed to detect and analyze phosphorylation of serine residues in proteins. It is a valuable tool for researchers studying cellular signaling pathways and protein modifications.
Automatically generated - may contain errors
2 protocols using anti phospho ser antibody
1
Phosphorylation of N-Terminal Pyrin
2
Phosphorylation of N-Terminal Pyrin
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Myc/his-tagged WT or S208A/S242A mutant N-terminal pyrin (aa’s 1- 330) was overexpressed into 293T cells, and the N-terminal pyrin proteins were purified using His-SpinTrap™ (28-4013-53, GE Healthcare). Each purified N-terminal pyrin protein was incubated with 5 μg of PKN1 (PR7255B) or PKN2 (PR7370A, Thermo Scientific) at 30°C for 0.5h in a kinase buffer (50 mM HEPES pH7.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA, 500 μM ATP and 2.5 mM DTT). The reactants were analyzed by immunoblotting with anti-phospho-Ser antibody (9606, Cell Signaling) or staining with Pro-Q diamond (P33300, Invitrogen). Pro-Q diamond-stained gel was visualized on Molecular Imager FX (Bio-Rad).
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