Purification, identification, and cloning of lysoplasmalogenase, the enzyme that catalyzes hydrolysis of the XXXX bond of lysoplasmalogen.

@entity1 @entity4990 @entity72706 @entity6072 @entity28053 @entity13475 @entity72705 @entity17046 @entity35 @entity15001 @entity526
Lysoplasmalogenase (EC 3.3.2.2 and EC 3.3.2.5) is an enzyme that catalyzes hydrolytic cleavage of the @entity17046 bond of lysoplasmalogen, forming @entity72705 and @entity28053 or @entity15001 and is specific for the sn-2-deacylated form of plasmalogen. Here we report the purification, characterization, identification, and cloning of lysoplasmalogenase. @entity35 liver microsomal lysoplasmalogenase was solubilized with @entity6072 and purified 500-fold to near homogeneity using four chromatography steps. The purified enzyme has apparent K(m) values of    50  m for both @entity13475 and @entity13475 and apparent V(m) values of 24.5 and 17.5  mol/min/mg protein for the two substrates, respectively. The pH optimum was 7.0. Lysoplasmalogenase was competitively inhibited by @entity4990 (K(i)    20  m). The predominant band on a gel at    19 kDa was subjected to trypsinolysis, and the peptides were identified by mass spectrometry as @entity72706 , a protein of unknown function. Transient transfection of @entity1 embryonic kidney (HEK) 293T cells showed that @entity72706 cDNA yielded lysoplasmalogenase activity, and Western blot analyses confirmed the synthesis of @entity72706 protein. The protein was localized in the membrane fractions. The @entity72706 gene was also transformed into @entity526 , and its expression was verified by Western blot and activity analyses. @entity72706 is a hydrophobic transmembrane protein of the YhhN family. Northern blot analyses demonstrated that liver expressed the highest level of @entity72706 , which agreed with tissue distribution of activity. Overexpression of @entity72706 in HEK 293T cells resulted in decreased levels of plasmalogens, suggesting that the enzyme may be important in regulating plasmalogen levels in animal cells.
