|Detection of Human PIKFyve by Western Blot. Western blot shows lysates of Jurkat human acute T cell leukemia cell line and K562 human chronic myelogenous leukemia cell line. PVDF membrane was probed with 0.25 µg/mL of Sheep Anti-Human PIKFyve Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7885) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for PIKFyve at approximately 240 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.|
PIKfyve (PhosphoInositide Kinase for five position containing a fyve finger (FAB1/YOTB/Vac1/EEA1) domain; also phosphatidylinositol 3-phosphate 5-kinase type III and Type III PIP kinase) is a 230-260 kDa cytoplasmic and endocytic member of an ancient family of phosphatidylinositol-3,5 biphosphate (PI[3,5]P2 synthesizing enzymes. It is associated with endosomes, particularly the microdomains of early endosomes that are EEA1-deficient. PIKfyve binds to membrane-embedded phosphatidylinositol and initially converts PI3P into PI(3,5)P2, which is then converted into PI5P. In conjunction with ArPIKfyve and Sac3, its actions are involved in the normal transport, fusion, and membrane export of endosomes. Notably, PIKfyve also appears to act as a protein kinase, apparently controlling its own activity through autophosphorylation. Human PIKfyve is 2098 amino acids (aa) in length. It contains one FYVE PI3P-binding zinc-finger domain (aa 154-219), a DEP domain (aa 365-440) and a C-terminal PIP kinase rehion (aa 1172-2085). There are no less than 29 utilized Ser/Thr phosphorylation sites, plus two utilized Tyr phosphorylation sites. Three potential isoform variants are reported, all which possess a three aa substitution for aa 546-2098. Two of these three also demonstrate a deletion of aa 108-204, and a nine aa substitution for aa 109-203, respectively. Over aa 215-361, human and mouse share 95% aa sequence identity.