Introduction
PHOSPHO1, involved in the mineralization process, plays a crucial role in bone and cartilage matrix mineralization. Its expression is evident at mineralization sites within these tissues, with high levels observed in the osteoblast cell line SaOS-2, known for producing a mineralized matrix. While Orphan-1, a collagen type-2, specifically targets cartilaginous tissues and is essential for normal skeletal embryonic development, linear growth, and cartilage's ability to withstand compressive forces, Phosphoethanolamine (2-O3POCH2CH2NH3) acts as a key intermediate in cephalin formation, occurring through ethanolamine phosphorylation in the liver and brain. PHOSPHO2 and PHOSPHO1, despite their similarities, exhibit subtle variations in charge distribution around the presumed substrate entry site and the positioning of potential H-bond donors. Distinguished by its high specific phosphoethanolamine and phosphocholine phosphatase activities, PHOSPHO1, a phosphatase enzyme with elevated expression in mineralizing cells, is involved in generating Pi for matrix mineralization - a critical process in skeletal development. As a member of the haloacid dehalogenase (HAD) superfamily of Mg2+-dependent hydrolases, PHOSPHO1 exhibits notable specific activities towards phosphoethanolamine (PEA) and phosphocholine (PCho).
Description
Recombinant Human Phospho1, produced in E.Coli, is a single, non-glycosylated polypeptide chain comprising 295 amino acids. With a molecular weight of 31.3 kDa, it features a 14 aa His tag at the N-Terminus. Purification of Human Phosphocholine Phosphatase is achieved through proprietary chromatographic techniques.
Physical Appearance
It appears as a filtered, lyophilized (freeze-dried) powder.
Formulation
The product is filtered (0.4µm) and lyophilized from a solution of 0.5mg/ml in 30mM acetate buffer with a pH of 4.
Solubility
To prepare a working stock solution of around 0.5mg/ml, add 0.1M Acetate buffer (pH 4) to the lyophilized pellet and allow it to dissolve completely. For conversion to a higher pH value, dilute the solution extensively with the appropriate buffer to a concentration of 10µg/ml. Note that the protein's solubility is limited at higher concentrations. The product is not sterile; filter it through a suitable sterile filter before using it in cell cultures.
Stability
Store the lyophilized protein at -20°C. After reconstitution, aliquot the product to minimize repeated freeze-thaw cycles. The reconstituted protein remains stable at 4°C for a limited duration.
Purity
The purity is determined to be greater than 95% through SDS-PAGE analysis.
Synonyms
Phosphoethanolamine/phosphocholine phosphatase, Phosphatase, Orphan 1, EC 3.1.3.75, Phospho1.
Amino Acid Sequence
MRGSHHHHHH GMASMSGCFP VSGLRCLSRD GRMAAQGAPR FLLTFDFDET IVDENSDDSI VRAAPGQRLP ESLRATYREG FYNEYMQRVF KYLGEQGVRP RDLSAIYEAI PLSPGMSDLL QFVAKQGACF EVILISDANT FGVESSLRAA GHHSLFRRIL SNPSGPDARG LLALRPFHTH SCARCPANMC KHKVLSDYLR ERAHDGVHFE RLFYVGDGAN DFCPMGLLAG GDVAFPRRGY PMHRLIQEAQ KAEPSSFRAS VVPWETAADV RLHLQQVLKSC.
