Apolipoprotein B-100 (apoB-100) appears in three forms in the endoplasmic reticulum of Hep G2 cells: (1) tightly bound to the membrane, ie, not extractable by sodium carbonate. This form is glycosylated but protease sensitive when present in intact microsomes, suggesting that it is only partially translocated to the microsomal lumen; (2) extractable by sodium carbonate and present on low-densit...

We have used the homobifunctional cross-linking reagent disuccinimidyl suberate (DSS) to identify proteins that are adjacent to nascent polypeptides undergoing translocations across mammalian rough ER. Translocation intermediates were assembled by supplementing cell free translations of truncated mRNAs with the signal recognition particle (SRP) and microsomal membrane vesicles. Two prominent cr...

Hydropathy profiles of 34 aligned cytochrome P-450 sequences were compared to identify potential transmembrane segments. Eleven regions with the potential to cross a membrane in at least some P-450 sequences were detected. The known sidedness of several residues and peptides was used to eliminate some of these regions from consideration. Further arguments based on the location and orientation o...

The precursor to the Euglena light-harvesting chlorophyll a/b-binding protein of photosystem II (pLHCPII) is unique; it is a polyprotein, synthesized on membrane-bound ribosomes and transported to the Golgi apparatus prior to chloroplast localization. A cDNA corresponding to the 5' end of LHCPII mRNA has been isolated and sequenced. The deduced amino acid sequence of this cDNA indicates that Eu...

We investigated the influence of pinoline (0.01-1.5 mM) on microsomal membrane fluidity before and after rigidity was induced by oxidative stress. In addition, we tested the effect of pinoline in the presence of 1 mM melatonin. The fluidity in rat hepatic microsomes was monitored using fluorescence spectroscopy and it was compared to the inhibition of malonaldehyde (MDA) plus 4-hydroxyalkenals ...

A rapid method of preparing plasma membranes from isolated fat cells is described. After homogenization of the cells, various fractions were isolated by differential centrifugation and linear gradients. Ficoll gradients were preferred because total preparation time was under 3 hr. The density of the plasma membranes was 1.14 in sucrose. The plasma membrane fraction was virtually uncontaminated ...

Rat liver microsomes (microsomal fraction) were isolated from vitamin A-deficient and -sufficient rats and analysed for membrane lipid characteristics. Membrane fluidity was found to be significantly decreased in microsomes from the vitamin A-deficient rats, but not in liposomes prepared from lipid extracts. Microsomes from vitamin A-deficient animals showed a significant decrease in C18:2, ome...

High affinity Ca2+-stimulated Mga+-dependent ATPase activity of nerve ending particles (!vnaphmm) from rat brain tissue appears to be associated primarily with isolated synaptic plasma membranes. The synaptic membrane (Ca2+ + Mga+)-ATPase activity was found to exhibit strict dependence on Mg+ for the presence of the activity, a high affinity for Ca2+ (&.a = 0.23 NM), and relatively high affini...

The presence of CYP2D6 at the surface of isolated rat and human hepatocytes and its recognition by autoantibodies were reported recently. We wondered whether the unexpected outside orientation at the plasma membrane could be related to topological inversion (luminal-oriented form) of cytochrome P450 in the endoplasmic reticulum. To examine the potential role of cDNA polymorphism, a CYP2D6 varia...

The NH2-terminal sequences of cytochromes P-450 resemble signal peptides, but these sequences are not cleaved during the insertion of these integral membrane proteins into the microsomes. To examine whether these putative signal peptides are functionally equivalent to signal peptides of secretory proteins, cDNA coding for a fusion protein was produced, in which the signal peptide for prepropara...