Alkaline phosphatase

Ball and stick model of alkaline phosphatase

Alkaline phosphatase (ALP) (EC 3.1.3.1) is a hydrolase enzyme responsible for removing phosphate groups from many types of molecules, including nucleotides, proteins, and alkaloids. The process of removing the phosphate group is called dephosphorylation. As the name suggests, alkaline phosphatases are most effective in an alkaline environment.

Bacterial

In bacteria, alkaline phosphatase is located in the periplasmic space, external to the cell membrane. Since this space is much more subject to environmental variation than the actual interior of the cell, bacterial alkaline phosphatase is comparatively resistant to inactivation, denaturation, and degradation, and also has a higher rate of activity. Although the actual purpose of the enzyme is still not fully understood, the simple hypothesis, that it is a means for the bacteria to generate free phosphate groups for uptake and use, is supported by the fact that alkaline phosphatase is usually only produced by the bacteria during phosphate starvation and not when phosphate is plentiful. However, other possibilities exist; for instance, the presence of phosphate groups usually prevents organic molecules from passing through the membrane, therefore dephosphorylating them may be important for bacterial uptake of organic compounds in the wild. Some complexities of bacterial regulation and metabolism suggest that other, more subtle, purposes for the enzyme may also play a role for the cell. In the laboratory, however, mutant Escherichia coli lacking alkaline phosphatase survive quite well, as do mutants unable to shut off alkaline phosphatase production.

Use in research

The most common alkaline phosphatases used in research are:

• Bacterial alkaline phosphatase (BAP), from Escherichia coli C4 cells
• Shrimp alkaline phosphatase (SAP), from a species of arctic shrimp (Pandalus borealis)
• Calf intestine alkaline phosphatase (CIAP), from calf intestine

Alkaline phosphatase has become a useful tool in molecular biology laboratories, since DNA normally possesses phosphate groups on the 5' end. Removing these phosphates prevents the DNA from ligating (the 5' end attaching to the 3' end of another molecule), thereby preventing DNA degradation until the next step of the process for which it is being prepared; also, removal of the phosphate groups allows radiolabeling (replacement by radioactive phosphate groups) in order to measure the presence of the labeled DNA through further steps in the process or experiment. For these purposes, the alkaline phosphatase from shrimp is the most useful, as it is the easiest to inactivate once it has done its job.

Another important use of alkaline phosphatase is as a label for enzyme immunoassays.

One common use in the dairy industry is as a marker of pasteurisation. This molecule is denatured by elevated temperatures found during pasteurisation, and can be tested for via colour change of a para-nitro-phenol phosphate substrate in a buffered solution. (Aschaffenburg Mullen Test)Raw milk would typically produce a yellow colouration within a couple of minutes, whereas properly pasteurised milk should show no change. There ARE of course exceptions to this in the case of heat stable alkaline phophatases produced by some bacteria.

Human

alkaline phosphatase, intestinal
Identifiers
Symbol	ALPI
NCBI gene	248
HGNC	437
OMIM	171740
RefSeq	NM_001631
UniProt	P09923
Other data
EC number	3.1.3.1
Locus	Chr. 2 q37.1
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alkaline phosphatase, liver/bone/kidney
Identifiers
Symbol	ALPL
Alt. symbols	HOPS
NCBI gene	249
HGNC	438
OMIM	171760
RefSeq	NM_000478
UniProt	P05186
Other data
EC number	3.1.3.1
Locus	Chr. 1 p36.12
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alkaline phosphatase, placental (Regan isozyme)
Identifiers
Symbol	ALPP
NCBI gene	250
HGNC	439
OMIM	171800
RefSeq	NM_001632
UniProt	P05187
Other data
EC number	3.1.3.1
Locus	Chr. 2 q37.1
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Physiology

In humans, alkaline phosphatase is present in all tissues throughout the entire body, but is particularly concentrated in liver, bile duct, kidney, bone, and the placenta. The optimal pH for the enzyme activity is pH=10 in standard conditions (310K,1 atm)

Diagnostic use

Concentrations blood plasma (serum) levels of ALP are typically 3-15 Units per liter, depending on the assay and local normal guidelines.

Lowered levels of ALP are less common than elevated levels.

The following conditions can cause abnormal levels of ALP:

Elevated levels (hyperphosphatasemia)

If it is unclear why alkaline phosphatase is elevated, isoenzyme studies using electrophoresis can confirm the source of the ALP. Heat stability also distinguishes bone and liver isoenzymes ("bone burns, liver lasts").

• Liver (Liver ALP):
  • Cholestasis, cholecystitis, cholangitis, cirrhosis, hepatitis, fatty liver, liver tumor, liver metastases, drug intoxication
  • Drugs: e.g. verapamil, carbamazepine, phenytoin, erythromycin, allopurinol, ranitidine
• Bone disease (Bone ALP):
  • Paget's disease, osteosarcoma, bone metastases of prostatic cancer (High / very high ALP values)
  • Other bone metastases
  • Fractured bone
  • Multiple myeloma (only when associated with fractures)
• Skeletal involvement of other primary diseases:
  • Osteomalacia, rickets, vitamin D deficiency, (Moderate rise)
  • Malignant tumors (ALP originating from tumor)
  • Renal disease (secondary hyperparathyroidism)
  • Primary hypothyroidism
• Polycythemia vera
• Myelofibrosis
• Leukemoid reaction to infection
• women on oral contraceptives
• Pregnancy
• Biliary obstruction
• Transient hyperphosphatasaemia of infancy: benign, often assocoated with infection

Lowered levels (hypophosphatasemia)

• Hypophosphatasia, an autosomal recessive disease
• Postmenopausal women receiving estrogen therapy because of osteoporosis
• Men with recent heart surgery, malnutrition, magnesium deficiency, hypothyroidism or severe anemia
• Children with achondroplasia and cretinism
• Children after a severe enteritis
• Pernicious anemia
• Aplastic anemia
• Chronic myelogenous leukemia

Other notes

Leukocyte alkaline phosphatase (LAP) is found within white blood cells.

See also

• Liver function tests

External links

• IUBMB Enzyme Nomenclature
• Structure and Mechanism of Alkaline Phosphatase