Measurement of alkaline phosphatase activity: characterization and identification of an inactivator in 2-amino-2-methyl-1-propanol.
An inactivator of alkaline phosphatase (EC 3.1.3.1) in 2-amino-2-methyl-1-propanol is demonstrated and characterized. This time-dependent inactivation results from chelation of enzyme-bound Zn2+; it is reversed by addition of Zn2+ and, to a lesser extent, other divalent metal ions. Cu2+ is an effective spectral indicator and can be used to determine the presence and quantity of inactivator. Data obtained from enzyme inactivation, Cu2+ absorbance spectra, "high-performance" liquid chromatography, thin-layer chromatography, Fourier-transform infrared spectroscopy, and mass spectroscopy indicate that the inactivator is 5-amino-3-aza-2,2,5-trimethylhexanol. This compound, even in trace amounts (less than 0.05% on a molar basis), shown to inactivate alkaline phosphatase.
D000469:Alkaline Phosphatase / Q000037:antagonists & inhibitors / Q000097:blood*; D002021:Buffers / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003300:Copper / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D011412:Propanolamines / Q000494:pharmacology*; D013053:Spectrophotometry / Q000379:methods; D015032:Zinc / Q000494:pharmacology
D002021:Buffers; D011412:Propanolamines; D003300:Copper; D000469:Alkaline Phosphatase; D015032:Zinc; C006551:2-amino-2-methyl-1-propanol