Leukotriene D4 and E4 formation by plasma membrane bound enzymes.
The homogenate of rat basophilic leukemia cells produces both the dihydroxy-leukotrienes and the peptido-leukotrienes (LT) C4, D4 and E4. The enzymes responsible for the formation of LTA4 and LTB4 are in the soluble fraction while the enzymes for LTC4, LTD4 and LTE4 are particulate (10,000 X g pellet). Centrifugation of the 10,000 X g pellet over a sucrose gradient resulted in two subfractions, a membrane fraction and a pellet (sucrose pellet). The fractions were incubated with LTC4, and the products were identified by bioassay, HPLC and UV spectra. The membrane fraction contained the enzymes gamma-glutamyl transpeptidase and amino peptidase which convert LTC4 to LTD4 and LTD4 to LTE4, respectively. When incubated with LTC4, the membrane fraction showed a dose dependent formation of LTD4 and a time course which reached a plateau at 30 to 45 minutes. Addition of serine.borate blocked the formation of LTD4, and cysteine blocked LTE4 production. The sucrose pellet showed little conversion of LTC4 to LTD4. We conclude that the gamma-glutamyl transpeptidase and the amino peptidase which produce LTD4 and LTE4 respectively are plasma membrane bound.
D000626:Aminopeptidases / Q000378:metabolism; D000818:Animals; D001491:Basophils / Q000201:enzymology; D002250:Carbon Radioisotopes; D002462:Cell Membrane / Q000201:enzymology / Q000648:ultrastructure; D002851:Chromatography, High Pressure Liquid; D007700:Kinetics; D007942:Leukemia, Experimental / Q000201:enzymology*; D017999:Leukotriene E4; D008854:Microscopy, Electron; D051381:Rats; D013189:SRS-A / Q000031:analogs & derivatives* / Q000096:biosynthesis*; D005723:gamma-Glutamyltransferase / Q000378:metabolism
D002250:Carbon Radioisotopes; D013189:SRS-A; D017999:Leukotriene E4; D005723:gamma-Glutamyltransferase; D000626:Aminopeptidases