On the spatial arrangememt of proteins in microsomal membranes from rat liver.
Rat liver rough microsomes were labeled enzymatically with (125)I using lactoperoxidase and glucose oxidase. In intact microsomes only proteins exposed on the outside face of the microsomal membrane were iodinated. Low concentrations of detergent (0.049% deoxycholate) were used to allow entrance of the iodination system into the vesicles without disassembling the membranes. This led to iodination of the soluble content proteins and to an increased labeling of the membrane proteins. The distribution of radioactivity in microsomal proteins was analyzed after separation by sodium dodecyl sulfate acrylamide gel electrophoresis. Most membrane proteins were labeled when intact microsomes were iodinated. No major membrane proteins were exclusively labeled in the presence of low detergent concentrations or after complete membrane disassembly. Therefore it is unlikely that there are major membrane proteins, other than glycoproteins, present only on the inner membrane face or completely embedded within the microsomal membrane. Microsomal proteins were also labeled by incubating rough microsomes with [(3)H]-NaBH(4) after reaction with pyridoxal phosphate. Microsomal membranes were permeable to these small molecular weight reagents as shown by the fact that proteins in the vesicular cavity as well as membrane proteins were labeled with this system.
D000818:Animals; D001894:Borohydrides; D002499:Centrifugation, Density Gradient; D002918:Chymotrypsin; D003840:Deoxycholic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D005949:Glucose Oxidase; D006651:Histocytochemistry; D007457:Iodine Radioisotopes; D008566:Membranes / Q000032:analysis; D008862:Microsomes, Liver / Q000032:analysis*; D010539:Permeability; D010544:Peroxidases; D011506:Proteins / Q000032:analysis*; D011732:Pyridoxal Phosphate; D051381:Rats; D012967:Sodium Dodecyl Sulfate; D013056:Spectrophotometry, Ultraviolet; D013501:Surface-Active Agents; D014316:Tritium; D014357:Trypsin
D001894:Borohydrides; D007457:Iodine Radioisotopes; D011506:Proteins; D013501:Surface-Active Agents; D003840:Deoxycholic Acid; D014316:Tritium; D012967:Sodium Dodecyl Sulfate; D011732:Pyridoxal Phosphate; D005949:Glucose Oxidase; D010544:Peroxidases; D002918:Chymotrypsin; D014357:Trypsin