Alkaline Phosphatase Chemical Properties
- Density 1.12 g/mL at 20 °C
- storage temp. 2-8°C
- form suspension
- color white
- Specific Gravity1
- Water Solubility It is soluble in water.
- Stability:Stability Incompatible with strong oxidizing agents
- EPA Substance Registry SystemAlkaline phosphatase (9001-78-9)
Alkaline Phosphatase Usage And Synthesis
- Chemical Propertiessolid
- UsesHydrolyzes 5′-terminal monophosphate (dephosphorylation) from DNA and RNA. Prevents fragments from self annealing. 5′-nucleic acid targeting for probes.
- UsesAlkaline Phosphatase is commonly used to remove the 5′-terminal phosphate from nucleic acids.
- Purification MethodsThe E.coli supernatant in sucrose (20%, 33mM) in Tris-HCl pH 8.0 is purified through a DEAE-cellulose column and recrystallised. To the column eluates in 0.125M NaCl is added MgCl2 (to 0.01M) and brought to 50% saturation in (NH4)2SO4 by adding the solid (0.20g/mL). The mixture is centrifuged to remove bubbles and is adjusted to pH 8.0 (with 2N NaOH). Saturated (NH4)2SO4 at pH 8.0 is added dropwise until the solution becomes faintly turbid (~61% saturation). It is set aside at ~25o for 1hour (turbidity will increase). The mixture is placed in an ice bath for several minutes when turbidity disappears and a clear solution is obtained. It is then placed in a large ice bath at 0o (~5L) and allowed to warm slowly to room temperature in a dark room whereby crystals are formed appearing as a silky sheen. The crystals are collected by centrifugation at 25o if necessary. The crystalline solutions are stable at ~25o for many months. They can be stored at 0o, but are unstable when frozen. Cysteine at 10-3M and thioglycolic acid at 10-4M are inhibitory. This is reversed on addition of Zn2+ ions. Many organic phosphates are good substrates for this phosphatase. [Molamy & Horecker Methods Enzymol 9 639 1966, Torriani et al. Methods Enzymol 12b 212 1968, Engstrom Biochim Biophys Acta 92 71 1964.] Alkaline phosphatase from rat osteosarcoma has been purified by Me2CO precipitation and chromatography on DEAE-cellulose, Sephacryl S-200, and hydroxylapatite. [Nair et al. Arch Biochem Biophys 254 18 1987.] Phosphoproteins (various). These are purified by adsorbing onto an iminodiacetic acid substituted agarose column to which are bound ferric ions. This chelate complex acts as a selective immobilised metal affinity adsorbent for phosphoproteins. [Muszyfiska et al. Biochemistry 25 6850 1986.]
Alkaline Phosphatase Preparation Products And Raw materials
- Raw materials1-Butanol
- Magnesium carbonate hydroxide FUCHSIN BASIC ALKALINEPHOSPHATASE,(EC184.108.40.206) ALKALINEPHOSPHATASE(ALP)(EC220.127.116.11) Cupric acetylacetonate Ferric acetylacetonate COBALT(II) ACETYLACETONATE 2,4-PENTANEDIONE, SILVER DERIVATIVE TERT-BUTYL ISOCYANIDE Tosylmethyl isocyanide N-BUTYLISOCYANIDE SALCOMINE TRIS(2,2,6,6-TETRAMETHYL-3,5-HEPTANEDIONATO)DYSPROSIUM(III) COBALT ETHYLENE DIAMINE CHLORIDE Tris(2,4-pentanedionato)chroMiuM(III) Aluminum acetylacetonate DICHLORO(ETHYLENEDIAMINE)PLATINUM(II) METHYL ISOCYANOACETATE
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