Basic introduction of Alkaline phosphatase

Alkaline phosphatase (ALP or AKP) is widely distributed in the human liver, bones, intestines, kidneys and placenta and other organizations by the liver to the bile out of an enzyme. This enzyme catalyzes the removal of the 5 & apos; phosphate group by the nucleic acid molecule, thereby converting the 5 & apos; -P terminus of the DNA or RNA fragment into the 5 & apos; -OH terminus. But it is not a single enzyme, but a group of isoenzymes. At present, ALP1, ALP2, ALP3, ALP4, ALP5 and ALP6 have been found. The first, second and sixth species were from the liver, the third from the bone cells, the fourth from the placenta and cancer cells, and the fifth from the small intestinal villi epithelium and fibroblasts.

Chemical characterization of Alkaline phosphatase

Belonging to homodimer protein, molecular weight of 56KDa. Each monomer is composed of 449 amino acids. The complete ALP molecule exhibits a typical α / β topology, and each monomer has an active center. The active center consists of Asp101-Ser102-Ala103, Arg166, Water molecules, three metal ions and their ligand amino acid composition. ALP is encoded by the phoA gene, which, like many secretory proteins, synthesizes a monomer precursor with a signal peptide at the amino terminus in the cytoplasm. The signal peptide leads the precursor to be transected across the intima and removed, resulting in homodimer formation.

Alkaline phosphatase is an enzyme capable of dephosphorylating the corresponding substrate, i.e. by removal of the phosphate groups on the substrate molecule by the hydrolysis of the phosphoric monoester, and the formation of phosphate ions and free hydroxyl groups, Nucleic acids, proteins, alkaloids and the like. The dephosphorylation process is known as dephosphorylation or dephosphorylation. Alkaline phosphatase is a kind of phosphatase, the role of phosphatase and the role of kinase is the opposite, kinase is phosphorylase, can use energy molecules, such as ATP, the phosphate group added to the corresponding substrate molecules. Alkaline phosphatase has the highest activity in alkaline environment, the optimum pH for Alkaline phosphatase from bacteria is 8.0, and for Alkaline phosphatase from cattle is 8.5.

Alkaline phosphatase is a zinc-containing glycoprotein, in the alkaline environment (optimum Ph is about 10) can hydrolyze a variety of natural and synthetic phosphate monoester compound substrate.

Methods for the determination of Alkaline phosphatase

There are many, China has applied a wide range of color for the benzene disodium colorimetric method, but now the application is more continuous detection method.

The principle is the use of p-nitrophenol phosphate as substrate, 2-amino-2-methyl-1-propanol or diethanolamine as phosphate acyl acceptor. In the alkaline environment, Alkaline phosphatase catalyzes the hydrolysis of 4-NPP to produce free p-nitrophenol, which is converted to yellow in alkaline solution. The Alkaline phosphatase activity units were calculated from the rate of increase of absorbance at 405 nm.

The normal range of Alkaline phosphatase

(Continuous monitoring method)

Female, 1-12 years old less than 500U / L; more than 15 years old, 40-150U / L;

Male, 1-12 years old less than 500U / L; 12-15 years old, less than 750U / L; more than 15 years old, 40-150U / L.

High values may indicate bile duct obstruction; low values are more common in children and pregnant women. High values suggest that active bone deposition may occur because Alkaline phosphatase is a by-product of osteoblastic activity (osteogenesis) (eg, in Peggy's Disease [the case of deformity osteitis] in).

In general, low values are rarer than high values. Alkaline phosphatase has a low clinical significance of the clinical generally high Alkaline phosphatase shows hepatobiliary system diseases. Low Alkaline phosphatase can also be seen in chronic nephritis, anemia, hypothyroidism and so on. In addition, the low Alkaline phosphatase in patients with liver disease is also more common friends, this time need attention, it is recommended to do further examination to identify the cause, so symptomatic treatment.

Clinical significance of Alkaline phosphatase

Clinical determination of Alkaline phosphatase is mainly used for bone, hepatobiliary system disease diagnosis and differential diagnosis, in particular the differential diagnosis of jaundice. For unexplained high Alkaline phosphatase serum levels, isoenzymes can be measured to aid in clarifying their organ source.

1. Physiological increase: children in the physiological skeletal development, Alkaline phosphatase activity than normal 1 to 2 times higher. In the growing period of adolescents, as well as pregnant women and high fat content of food can be increased.

2. Pathological elevation:

(1) skeletal diseases such as rickets, rickets, bone malignancies, malignant bone metastasis;

(2) hepatobiliary diseases such as extrahepatic biliary obstruction, liver cancer, liver cirrhosis, capillary cholangiocarcinoma;

(3) other diseases such as hyperparathyroidism.

3. Pathological reduction: found in severe chronic nephritis, thyroid dysfunction in children, anemia.

Alkaline phosphatase inhibition

All Alkaline phosphatase isozymes from all mammals except the placenta can be inhibited by high arginine; all Alkaline phosphatase isozymes except intestinal and placental can be inhibited by levamisole; and in addition to from the placenta Of Alkaline phosphatase SEAP, almost all Alkaline phosphatase can be inactivated at 2 hours at 65 ° C.

Monoethanolamine has a temperature-dependent inhibitory effect on Alkaline phosphatase;

Higher concentrations of inorganic phosphorus competitively inhibited the activity of Alkaline phosphatase;

EDTA can cause irreversible inhibition of Alkaline phosphatase activity by complexing with Alkaline phosphatase, leading to the change of microenvironment conformation of the enzyme active center.

Other inhibitors include benzidine (competitive), L-phenylalanine and dansyl-L-phenylalanine (anti-competitive), nitrogen-containing heterocyclic vanadyl complexes (selectivity), pentachloro Phenol (anti-competitive inhibition of human placental Alkaline phosphatase) and the like.

Human Sources of Alkaline phosphatase and Its Application

The human condition: Alkaline phosphatase is mainly used for obstructive jaundice, primary liver cancer, secondary liver cancer, cholestatic hepatitis and other checks. When suffering from these diseases, excessive production of liver cells ALKALINE PHOSPHATASE, the lymphatic and hepatic sinus into the blood, while the bile due to intrahepatic bile excretion obstacles, anti-inflow of blood caused by serum Alkaline phosphatase was significantly increased. However, this enzyme is also active in bone tissue. Therefore, pregnant women, fracture healing, osteomalacia. Rickets, bone cell cancer, osteoporosis, liver abscess, liver tuberculosis, cirrhosis, leukemia, hyperthyroidism, serum Alkaline phosphatase can also be increased, should be identified.

Research application: Alkaline phosphatase is also one of the most commonly used labeling reagents in immunodiagnostic reagents. Compared with HRP, Alkaline phosphatase has the advantages of high stability, high sensitivity, and high cost. It is difficult to label.