Hepatitis C virus antibodies among risk groups in Spain.

Activation of pro-caspase-3 is a central event in the execution phase of apoptosis and appears to serve as the convergence point of different apoptotic signaling pathways. Recently, mitochondria were found to play a central role in apoptosis through release of cytochrome c and activation of caspases. Moreover, a sub-population of pro-caspase-3 has been found to be localized to this organelle. In the present study, we demonstrate that pro-caspase-3 is present in the mitochondrial fraction of Jurkat T cells in a complex with the chaperone proteins Hsp60 and Hsp10. Induction of apoptosis with staurosporine led to the activation of mitochondrial pro-caspase-3 and its dissociation from the Hsps which were released from mitochondria. The release of Hsps occurred simultaneously with the release of other mitochondrial intermembrane space proteins including cytochrome c and adenylate kinase, prior to a loss of mitochondrial transmembrane potential. In in vitro systems, recombinant Hsp60 and Hsp10 accelerated the activation of pro-caspase-3 by cytochrome c and dATP in an ATP-dependent manner, consistent with their function as chaperones. This finding suggests that the release of mitochondrial Hsps may also accelerate caspase activation in the cytoplasm of intact cells.

/pdf/presence-of-a-pre-apoptotic-complex-of-pro-caspase-3-hsp60-1atzzfbylx.pdf

Presence of a pre-apoptotic complex of pro-caspase-3, Hsp60 and Hsp10 in the mitochondrial fraction of jurkat cells.

DT-diaphorase as a quinone reductase: a cellular control device against semiquinone and superoxide radical formation.

Abbreviations: 
ACD, Annihilation coincidence detection; BGO, bismuth germanate; CBF, cerebral blood flow; CBV, cerebral blood volume; CMG, 3-(11C)-methyl-D-glucose; CMRGlc, cerebral metabolic rate for glucose; CMR02, cerebral metabolic rate for oxygen; CsF, cesium fluoride; CT, computed tomography; DG, deoxyglucose; FDG, 2-fluoro-2-deoxy-d-glucose; LC, lumped constant; OEF, oxygen extraction fraction; PCT, positron computed tomography; PETT, positron emission transaxial tomography; PT, physiological tomography.

Study of Cerebral Function with Positron Computed Tomography

Since the pioneering work on structure and function of heteroglycans compiled in the classical books edited by A. Gottschalk in 19721, there have been several promising developments in glycoconjugate research, as reviewed in this article.

/pdf/structure-biosynthesis-and-functions-of-glycoprotein-glycans-43rkhpv8hp.pdf

Structure, biosynthesis and functions of glycoprotein glycans

1
Protease and diazobenzene sulfonate have been used to probe the transverse topology of the microsomal glucose-6-phosphatase system.

2
Treatment of intact microsomes with these reagents under the conditions used here did not affect the permeability of the membrane to mannose 6-phosphate, nucleoside diphosphatase, or dextran of 70 000 molecular weight. Nor did these treatments inactivate the hydrolytic site of glucose-6-phosphatase, a finding in agreement with earlier conclusions that this site is on the inside of the membrane.

3
On the other hand, treatment of intact microsomes with diazobenzene sulfonate or proteases does inactivate (or increase the apparent Km of) some other component which is rate-limiting for glucose-6-phosphatase activity in intact but not in disrupted microsomes. The simplest explanation for this phenomenon is that there is a protein carrier in the microsomal membrane which transports glucose 6-phosphate from the medium to the lumen, where it is hydrolyzed, and that diazobenzene sulfonate and proteases attack this carrier.

4
The lack of effect of these reagents on microsomal inorganic pyrophosphatase activity suggests that the glucose-6-phosphate carrier cannot transport pyrophosphate.

5
Finally, it was found that treatment of microsomes with ammonia breaks down their permeability barrier but also removes significant amounts of microsomal phospholipid and inactivates a number of microsomal enzymes. We do not recommend it as a general approach to altering microsomal permeability.

Evidence for the Involvement of a Glucose-6-Phosphate Carrier in Microsomal Glucose-6-Phosphatase Activity

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/0014-5793%2875%2980256-0

Distribution of constitutive enzymes and phospholipids in microsomal membranes of rat liver

Investigation of the transverse topology of the microsomal membrane using combinations of proteases and the non-penetrating reagent diazobenzene sulfonate.

Mannosylation of the proteins of microsomal and Golgi membranes was investigated both after incubation in vitro of the isolated subfractions with GDP-[14C]mannose and after injection of [3H]mannose into rats followed by separation of these subfractions. Mannosylation of endogenous and added exogenous dolichol phosphate and also of dolichol pyrophosphate-oligosaccharide occurs in all three fractions. It was essential to inhibit antagonistic enzymes during incubation and to centrifuge after incubation. The presence of detergent in the incubation mixture influences the incorporation pattern of the different fractions in very different ways. In a system in vitro predominantly membrane proteins and not secretory proteins are mannosylated. Trypsin treatment of intact vesicles removes components from the outer surface only; such treatment liberates about one third of the radioactive mannose associated with lipid, releases radioactivity from the protein acceptor to the same extent and causes some inactivation of the transferase activities. It appears that a part of the mannosyl transferase system in rough and smooth endoplasmic reticulum and in Golgi membranes is localized at the cytoplasmic side of these membranes. This activity is probably involved in the glycosylation of proteins localized at the cytoplasmic surface of the endoplasmic reticulum.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1978.tb12566.x

Mannosylation of Endogenous Proteins of Rough and Smooth Endoplasmic Reticulum and of Golgi Membranes

High doses of the lysosomotropic drug chloroquine result in lipid storage in many organs in animals. We used miniature pigs, type Gottingen, to study the lipid accumulation in skeletal muscle after chloroquine intoxication for more than 200 days. The lipids of the quadriceps muscle in intoxicated and in age-matched control pigs were characterized and determined. The lipid storage was larger in skeletal muscle than in any other organ of the intoxicated pigs. The concentration of phospholipids was increased threefold, acidic phospholipids relatively more than neutral ones. The lysosome-specific acidic phospholipid bis(monoacylglyceryl)phosphate content was almost 50-fold larger in the intoxicated pigs than in the controls. Cholesterol was increased slightly more than the phospholipids, but there was no particular accumulation of cholesteryl esters, which has been shown to occur in the liver. For the first time a storage of gangliosides, relatively more pronounced than of other lipids, was demonstrated in skeletal muscle in drug-induced lipidosis. The concentration of total gangliosides was increased 10–15-fold, and the pattern of gangliosides showed some distinct changes resulting in at least a 100-fold increase in the concentration of ganglioside GM2 (II3NeuAc-GgOse4Cer).

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1981.tb05373.x

Olle S. Nilsson

Papers

Evidence for the Involvement of a Glucose-6-Phosphate Carrier in Microsomal Glucose-6-Phosphatase Activity

Distribution of constitutive enzymes and phospholipids in microsomal membranes of rat liver

Investigation of the transverse topology of the microsomal membrane using combinations of proteases and the non-penetrating reagent diazobenzene sulfonate.

Mannosylation of Endogenous Proteins of Rough and Smooth Endoplasmic Reticulum and of Golgi Membranes

Chloroquine‐Induced Accumulation of Gangliosides and Phospholipids in Skeletal Muscles