Two sensitive, convenient, and widely applicable assays for marker enzyme activities specific to endoplasmic reticulum.
Two assay protocols are described for enzyme activities known to reside in the endoplasmic reticulum of a wide variety of species and tissue types, with the intent that they be used as marker enzyme assays in subcellular fractionations. The enzyme activities assayed are choline phosphotransferase and dolichol-P-mannosyl synthase, both of which result in synthesis of lipid products. The assays are constructed to make them easy to perform and sensitive enough to detect enzyme activity even using microgram quantities of cell protein. The assay methodologies are effective not only in vertebrate cells, but in insect cells and yeast cells as well. This implies that these assays should be useful as marker enzyme assays for a wide variety of eukaryotic cells.
Braell
W A
WA
Department of Biological Chemistry, Harvard Medical School, Boston, Massachusetts 02115.
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D002499:Centrifugation, Density Gradient; D002798:Diacylglycerol Cholinephosphotransferase / Q000302:isolation & purification*; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D006602:Hexosyltransferases / Q000302:isolation & purification*; D007313:Insecta / Q000201:enzymology; D008364:Mannosyltransferases / Q000302:isolation & purification*; D010770:Phosphotransferases / Q000302:isolation & purification*; D013045:Species Specificity; D013347:Subcellular Fractions / Q000201:enzymology; D015003:Yeasts / Q000201:enzymology
D006602:Hexosyltransferases; D008364:Mannosyltransferases; C022755:dolichyl-phosphate beta-D-mannosyltransferase; D010770:Phosphotransferases; D002798:Diacylglycerol Cholinephosphotransferase