Specific localization and quantification of biotin transport components in yeast by use of a biotin-conjugated, impermeant, electron-dense label.
Two approaches are described for the localization and quantification of biotin transport components in yeast cells. One approach is based on tracing the fate of a radioactive affinity label for the biotin transport system, [14C]biotinyl-p-nitrophenyl ester (pBNP), through various stages of subcellular fractionations. A complementary method involves the use of a biotin-derivatized, impermeant, electron-dense, affinity-cytochemical label (ferritin-biotin conjugates) for subsequent visualization by electron microscopy. Values of approximately 8,000 and 4,000 sites/cell, respectively, were achieved by the two methods. Complicating factors, future perspectives and the relevance of the two methods to the isolation of transport components are discussed.
D000345:Affinity Labels; D001692:Biological Transport; D001710:Biotin / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D005293:Ferritins; D026901:Membrane Transport Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
D000345:Affinity Labels; D026901:Membrane Transport Proteins; D001710:Biotin; D005293:Ferritins