Recombinant human hyaluronan synthase 3 is phosphorylated in mammalian cells

摘要: Hyaluronan is a ubiquitous component of vertebrate extracellular and cell-associated matrices that serves as key structural skin, cartilage, eyes joints, plays important roles in dynamic cellular processes, including embryogenesis, inflammation, wound healing metastasis. synthesized by three homologous hyaluronan synthases designated HAS1, HAS2 HAS3 differ their tissue distribution, regulation enzymatic characteristics. Some progress has been made characterizing HAS transcripts distinguishing the properties various isoforms, but essentially nothing known about possible posttranslational modification. Using [32P]Pi radiolabelling recombinant FLAG (DYKDDDDK) epitope-tagged version human expressed COS-7 cells, we show serine-phosphorylated this phosphorylation can be enhanced number effectors – most significantly membrane-permeable analogue cAMP. By employing novel FLAG-tagged phosphorylated reference protein derived from EGFP (enhanced green fluorescent protein), were able to estimate stoichiometry FLAG–HAS3 phosphorylation. It was approx. 0.11 unstimulated cells increased much 0.32 stimulated with 8-(4-chlorophenylthio)-cAMP.