Alkaline phosphatase

[y_darkness]

Hi anybody know why, unlike most other enzymes alkaline phosphatase is not inactivated by SDS?

[admin]

Hello Darkness,
do you mean during western blot development or during SDS PAGE Electrophoresis?

During western blot development I believe that it may be less affected as the SDS is usually washed off by then during the blot washing and blocking.

I am not sure about SDS-PAGE in gel phosphatase activity however.

[danfive]

From wikipedia
AP from bacteria--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.

AP from mammalian sources
Article: Size and stability to sodium dodecyl sulfate of alkaline phosphatases from their three established human genes --> http://www.ncbi.nlm.nih.gov/pubmed/6487640

From the abstract:
Stability of the catalytic activity of the purified enzymes to SDS varied and was very dependent on pH. SDS at 1% (w/v) rapidly denatured both native and desialylated alkaline phosphatase from placenta at pH 7.5 but had little effect on these at pH 10.3. Compared with placenta, the native enzyme from liver had greater stability at pH 7.5 and both native and desialylated forms had lower stability at pH 10.3. The enzyme from intestinal mucosa was sharply different from the other two isoenzymes: SDS had little effect at pH 7.5 but very rapidly denatured the enzyme at pH 10.3. The size of alkaline phosphatases and their stability to SDS can be used to identify gene products and to recognize heterodimers formed between products of more than one gene.

Its robustness may be due to its large size, glycoprotein nature and unique structure--its a metalloenzyme with 2 Zn ions and 1 Mg ion closely spaced together. See Structure and mechanism of alkaline phosphatase --> http://www.ncbi.nlm.nih.gov/pubmed/1...?dopt=Abstract