P6774

Fosfatasi, alcalina

buffered aqueous solution, ≥2,000 DEA units/mg protein

• Sinonimo/i: Fosfatasi alcalina, Fosfoidrolasi di monoesteri ortofosforici (alcalino ottimale)
• Numero CAS: 9001-78-9
• UNSPSC Code: 12352204
• eCl@ss: 42010105
• NACRES: NA.54
• Numero CE: 3.1.3.1 (BRENDA, IUBMB)
• MDL number: MFCD00131849
• Specific activity: ≥2,000 DEA units/mg protein
• Biological source: bovine intestinal mucosa
• Concentration: 5-20 mg/mL

Proprietà

• biological source: bovine intestinal mucosa
• Quality Segment: 200
• form: buffered aqueous solution
• specific activity: ≥2,000 DEA units/mg protein
• mol wt: dimer ~160 kDa
• concentration: 5-20 mg/mL
• storage temp.: 2-8°C

Descrizione

General description

La fosfatasi alcalina intestinale bovina è una glicoproteina dimerica derivata dalla membrana. Ne esistono almeno tre isoforme, che generalmente possiedono due glicani N-legati e uno o più glicani O-legati per monomero.

La fosfatasi alcalina intestinale bovina è una glicoproteina dimerica derivata dalla membrana. Ne esistono almeno tre isoforme, che generalmente possiedono due glicani N-legati e uno o più glicani O-legati per monomero.2 L′enzima richiede zinco e ioni bivalenti di magnesio o calcio per svolgere l′attività.

Bovine intestinal alkaline phosphatase is a dimeric, membrane-derived glycoprotein. At least three isoforms exist, which typically possess two N-linked and one or more O-linked glycans per monomer.

Application

Grado di attività altamente specifico consigliato per la coniugazione di anticorpi e proteine.

La fosfatasi alcalina può essere utilizzata per defosforilare la caseina e altre proteine. La fosfatasi alcalina può essere adoperata anche per defosforilare i terminali 5′ del DNA o dell′RNA per prevenire l′auto-legatura. Il DNA e l′RNA possono anche essere marcati con il fosfato radiomarcato (tramite la T4 polinucleotide chinasi) dopo la defosforilazione con la fosfatasi alcalina.

Alkaline phosphatase is used for conjugation to antibodies and other proteins for ELISA, Western blotting, and histochemical detection. It is routinely used to dephosphorylate proteins, such as casein, and nucleic acids. It may be used for protein labeling when high sensitivity is required. Alkaline phosphatase may be also be used to dephosphorylate the 5′-termini of DNA or RNA to prevent self-ligation. DNA or RNA can also be tagged with radiolabeled phosphate (via T4 polynucleotide kinase) after dephosphorylation with alkaline phosphatase.. Product P6774 has been used during preparation of brain lysate[1].

The enzyme from Sigma has been used in the preparation of AP-BGG (alkaline phosphatase-bovine gamma globulin) conjugates.[2] It has been conjugated to sheep anti-rabbit IgG and sheep anti-rabbit IgM in immunological assays.[3] It has also been used to prepare double-enzyme conjugates with HRP for the simultaneous detection of three different intracellular Ig determinants in a single tissue section. [4]

Biochem/physiol Actions

L′enzima ha un′ampia specificità per gli esteri fosforici di alcoli, ammine, pirofosfato e fenoli. Viene utilizzato abitualmente per defosforilare le proteine e gli acidi nucleici.

Alkaline phosphatase, from bovine intestinal mucosa, is most stable in the pH range 7.5-9.5. The enzyme has a broad specificity for phosphate esters of alcohols, amines, pyrophosphate, and phenols and it requires zinc, and magnesium or calcium divalent ions for activity.

The enzyme is a glycoprotein containing approximately 12% carbohydrate (6% hexoses and 6% other neutral sugars). Each molecule of alkaline phosphatase contains four zinc atoms and four disulfide bridges.[5] Maximal activity with alkaline phosphatase is achieved in the presence of magnesium.[6] It catalyzes the hydrolysis of phosphate monoesters such as p-nitrophenyl phosphate, phenyl phosphate, phenolphthalein phosphate, α-glycerol phosphate, β-glycerol phosphate, 2-phosphorylglycerate, triosephosphate, glucose-6-phosphate, glucose 1-phosphate, fructose 1-phosphate, fructose 6-phosphate, adenosine 5-phosphate adenosine 3-phosphate, phosphoenolpyruvate, and β-nicotinamide adenine dinucleotide phosphate.[7][8][9] Arsenate, cysteine, iodine, inorganic phosphate, pyrophosphate, diisopropyl phosphate, triphenylphosphate, diisopropyl fluorophosphate, and L-phenylalanine are some of the strong inhibitors of alkaline phosphatase.[7][10][11]

Physical form

Solution in 3.0 M NaCl containing 5 mM MgCl₂, 0.2 mM ZnCl₂, and 30 mM triethanolamine, pH 7.6

Preparation Note

Affinity purified

Analysis Note

Le dimensioni delle confezioni sono basate su unità DEA.

Protein determined by biuret.

Other Notes

Un′unità DEA idrolizza 1 μmole di 4-nitrofenil fosfato in un minuto a pH 9,8 e a 37 °C (un′unità di glicina è equivalente a circa 3 unità DEA).

Informazioni sulla sicurezza

• pictograms: GHS08
• signalword: Danger
• hcodes: H334
• pcodes: P261 - P284 - P501
• Hazard Classifications: Resp. Sens. 1
• Classe di stoccaggio: 10 - Combustible liquids
• flash_point_f: Not applicable
• flash_point_c: Not applicable
• ppe: Eyeshields, Gloves, multi-purpose combination respirator cartridge (US)
• wgk: WGK 1