||Fleshood, H. L.; Pitot, H. C.
||The metabolism of O-phosphorylethanolamine in animal tissues. I. O- phosphorylethanolamine phospho-lyase: partial purification and characterization
||J Biol Chem
||The enzyme 0-phosphorylethanolamine phospho-lyase has been purified 107-fold from rabbit liver and 580-fold
from rat liver with the use of DEAE-cellulose and brushite chromatography. The pH optimum of the enzyme’s activity is 7.8 and enzyme preparations are unstable at temperatures above 55°C.
The apparent K, for 0-phosphorylethanolamine was found to be 6.1 X 10e4. Dialyzed preparations after treatment
with hydroxylamine were found to be almost completely dependent on the addition of pyridoxal phosphate. The Km
for this cofactor was 2.7 X l0-7 M. Inorganic phosphate was found to be a competitive inhibitor with an apparent Ki of 1.3 X 1O-3 M.
None of the cations or anions tested was found to be essential for enzyme activity or to have an ability to activate the enzyme. The enzyme was inactivated by heavy metal cations, sulfate ions, and cyanide ions. Inhibition of pchloromercuribenzoate could be reversed by dithiothreitol. Substrate specificity was tested with a variety of compounds; ethanolamine, ethylphosphate, ethylamine, ethylenimine, and 2-aminoethylphosphonate were not substrates for the enzyme.
The approximate molecular weight as estimated by Sephadex G-200 chromatography was 168,000. An Arrhenius
plot of enzyme activity showed a discontinuity with the energy of activation being 18,700 calories below 14°C and 10,700 calories at higher temperature. By means of 0-phosphorylethanolamine labeled with 14C in the amino carbon, it was shown by oxidation and Schmidt degradation of the enzymatically formed acetaldehyde that
the l4C was solely in the carbonyl carbon of the product. These data serve to confirm the mechanism of this reaction.