Activation of Progelatinase B by Membranes of Human Polymorphonuclear Granulocytes

2000 ◽  
Vol 381 (1) ◽  
pp. 49-55 ◽  
Author(s):  
Hansjörg Kolkenbrock ◽  
Jürgen Zimmermann ◽  
Gerd-R. Burmester ◽  
Norbert Ulbrich
Keyword(s):  
Plasma Membranes ◽  
Complete Removal ◽  
Cathepsin G ◽  
Serine Proteinases ◽  
Physiological Importance ◽  
Polymorphonuclear Granulocytes ◽  
Weak Binding ◽  
Human Granulocytes ◽  
Membrane Bound

AbstractIsolated human granulocyte plasma membranes contain progelatinase B. The binding of progelatinase B to the membrane, however, is relatively weak, and a considerable part of progelatinase B can be removed by simply washing the membrane with buffer. This detachment does not depend on the ionic strength of the buffer, indicating that electrostatic forces do not play an important role in the binding of progelatinase B to the membrane. A complete removal of progelatinase B is achieved by chromatography of neutrophil membranes on gelatin-agarose.The plasma membrane of human granulocytes activates added progelatinase B. This activation is inhibited by soybean trypsin inhibitor and is thus performed by membrane bound serine proteinases. In contrast to other reports that claimed an important role of elastase in activating progelatinase B, we found that this activation is mostly inhibited by chymostatin and not by elastatinal and is thus primarily due to cathepsin G. Proteinase 3 was shown to activate progelatinase B as efficient as neutrophil elastase, i. e. much weaker than cathepsin G. Binding of cathepsin G and elastase to the neutrophil membrane does not change their ability to activate progelatinase B. However, cathepsin G , the most potent activator of the three neutrophil serine proteinases, is only a weak activator, when compared to stromelysin-1. This, as well as only a weak binding of progelatinase B, make it doubtful that activation of membrane-bound progelatinase B by membrane-bound serine proteinases is of significant physiological importance.

scholarly journals Levels of Human Erythrocyte Membrane-Bound and Cytosolic Glycohydrolases Are Associated with Oxidative Stress in Erectile Dysfunction Patients

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
L. Massaccesi ◽  
G. V. Melzi d’Eril ◽  
G. M. Colpi ◽  
G. Tettamanti ◽  
G. Goi ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Plasma Membrane ◽  
Erectile Dysfunction ◽  
Plasma Membranes ◽  
Lag Time ◽  
Specific Marker ◽  
Human Erythrocyte Membrane ◽  
Key Enzyme ◽  
Membrane Bound

Oxidative stress (OS) and production of NO, by endothelium nitric oxide synthetase (eNOS), are involved in the pathophysiology of erectile dysfunction (ED). Moreover, OS induces modifications of the physicochemical properties of erythrocyte (RBC) plasma membranes and of the enzyme content of the same membranes. Due to their role in signalling early membrane alterations in OS-related pathologies, several plasma membrane and cytosolic glycohydrolases of human RBC have been proposed as new markers of cellular OS. In RBC, NOS can be activated and deactivated by phosphorylation/glycosylation. In this regulatory mechanism O-β-N-AcetylGlucosaminidase is a key enzyme. Cellular levels of O-GlcNAcylated proteins are related to OS; consequently dysfunctional eNOS O-GlcNAcylation seems to have a crucial role in ED. To elucidate the possible association between RBC glycohydrolases and OS, plasma hydroperoxides and antioxidant total defenses (Lag-time), cytosolic O-β-N-AcetylGlucosaminidase, cytosolic and membrane Hexosaminidase, membraneβ-D-Glucuronidase, andα-D-Glucosidase have been studied in 39 ED patients and 30 controls. In ED subjects hydroperoxides and plasma membrane glycohydrolases activities are significantly increased whereas Lag-time values and cytosolic glycohydrolases activities are significantly decreased. These data confirm the strong OS status in ED patients, the role of the studied glycohydrolases as early OS biomarker and suggest their possible use as specific marker of ED patients, particularly in those undergoing nutritional/pharmacological antioxidant therapy.

scholarly journals Detection of acyl-CoA-binding protein in human red blood cells and investigation of its role in membrane phospholipid renewal

1995 ◽  
Vol 306 (3) ◽  
pp. 793-799 ◽  
Author(s):  
H Fyrst ◽  
J Knudsen ◽  
M A Schott ◽  
B H Lubin ◽  
F A Kuypers
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Human Liver ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Human Red Blood Cells ◽  
Low Concentrations ◽  
Membrane Bound ◽  
High Concentrations

Acyl-CoA-binding protein (ACBP) has been identified in a number of tissues and shown to affect the intracellular distribution and utilization of acyl-CoA. We have detected ACBP in the cytosol but not the membrane of human red blood cells and, using an e.l.i.s.a. with antibodies prepared against human liver ACBP, found that its concentration was 0.5 microM. To investigate the role of ACBP in human red blood cells, we added purified human liver ACBP and radiolabelled acyl-CoA to isolated membranes from these cells. ACBP prevented high concentrations of acyl-CoA from binding to the membrane but could not keep the acyl-CoA in the aqueous phase at low concentrations. This suggested the presence of a pool in the membrane with a binding affinity for acyl-CoA that was greater than that of ACBP for acyl-CoA. In the presence of lysophospholipid, this membrane-bound pool of acyl-CoA was rapidly used as a substrate by acyl-CoA:lysophospholipid acyltransferase (LAT) to generate phospholipid from lysophospholipid. We also found that ACBP-bound acyl-CoA was preferred over free acyl-CoA as a substrate by LAT. These results are the first documentation that human red blood cells contain ACBP and that this protein can affect the utilization of acyl-CoA in plasma membranes of these cells. The interactions between acyl-CoA, ACBP and the membrane suggest that there are several pools of acyl-CoA in the human red blood cell and that ACBP may have a role in regulating their distribution and fate.

scholarly journals Status of Asp29 and Asp40 in the Interaction of Naja atra Cardiotoxins with Lipid Bilayers

Toxins ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 262 ◽  
Author(s):  
Guan-Lin Wu ◽  
Yi-Jun Shi ◽  
Chia-Hui Huang ◽  
Yuan-Chin Lee ◽  
Liang-Jun Wang ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Plasma Membranes ◽  
Structural Transition ◽  
Lipid Vesicles ◽  
Structural Flexibility ◽  
Structural Constraints ◽  
Ionization Time ◽  
Membrane Bound ◽  
Naja Atra

It is widely accepted that snake venom cardiotoxins (CTXs) target the plasma membranes of cells. In the present study, we investigated the role of Asp residues in the interaction of Naja atra cardiotoxin 1 (CTX1) and cardiotoxin 3 (CTX3) with phospholipid bilayers using chemical modification. CTX1 contains three Asp residues at positions 29, 40, and 57; CTX3 contains two Asp residues at positions 40 and 57. Compared to Asp29 and Asp40, Asp57 was sparingly modified with semi-carbazide, as revealed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass and mass/mass analyses. Thus, semi-carbazide-modified CTX1 (SEM-CTX1) mainly contained modified Asp29 and Asp40, while SEM-CTX3 contained modified Asp40. Compared to that of native toxins, trifluoroethanol easily induced structural transition of SEM-CTX1 and SEM-CTX3, suggesting that the structural flexibility of CTXs was constrained by Asp40. Modification of Asp29 and Asp40 markedly promoted the ability of CTX1 to induce permeability of cell membranes and lipid vesicles; CTX3 and SEM-CTX3 showed similar membrane-damaging activity. Modification of Asp residues did not affect the membrane-binding capability of CTXs. Circular dichroism spectra of SEM-CTX3 and CTX3 were similar, while the gross conformation of SEM-CTX1 was distinct from that of CTX1. The interaction of CTX1 with membrane was distinctly changed by Asp modification. Collectively, our data suggest that Asp29 of CTX1 suppresses the optimization of membrane-bound conformation to a fully active state and that the function of Asp40 in the structural constraints of CTX1 and CTX3 is not important for the manifestation of membrane-perturbing activity.

Vitamin E and ATPases: Protection of ATPase Activities by Vitamin E Supplementation in Various Tissues of Hypercholesterolemic Rats

2000 ◽  
Vol 70 (1) ◽  
pp. 3-7 ◽  
Author(s):  
Evin Ademoglu ◽  
Cahide Gökkusu ◽  
Sükrü Palandüz
Keyword(s):  
Vitamin E ◽  
Plasma Membranes ◽  
Cholesterol Content ◽  
Kidney Cortex ◽  
Hypercholesterolemic Diet ◽  
Membrane Atpase ◽  
Membrane Bound ◽  
Vitamin E Supplementation

It has been shown that the lipid composition of plasma membrane can be modified in vivo by dietary fat. It has also been observed that an increase in the cholesterol content of plasma membranes results in decreased activities of ATPases. In the present study, we evaluated the changes in the activities of ATPases from erythrocytes, hepatocytes, and kidney cortex caused by cholesterol-rich diet in rats and subsequently examined the role of vitamin E administration on the cholesterol-induced effects in these tissues. Administration of hypercholesterolemic diet to the rats for 4.5 months, significantly decreased membrane Na+-K+-ATPase and Ca+2-ATPase activities in comparison to the controls in all tissues studied. Vitamin E supplementation to the hypercholesterolemic rats led to a recovery in membrane ATPase activities. In conclusion, vitamin E supplementation to the rats provided protection against hypercholesterolemic diet-induced impairment of membrane-bound ATPases.

scholarly journals A membrane-bound activity catalysing phosphatidylinositol breakdown to 1,2-diacylglycerol, d-myoinositol 1:2-cyclic phosphate and d-myoinositol 1-phosphate. Properties and subcellular distribution in rat cerebral cortex

1973 ◽  
Vol 131 (3) ◽  
pp. 433-442 ◽  
Author(s):  
Eduardo G. Lapetina ◽  
Robert H. Michell
Keyword(s):  
Cerebral Cortex ◽  
Plasma Membranes ◽  
Surface Membrane ◽  
Rat Cerebral Cortex ◽  
Synaptic Membrane ◽  
First Order Kinetics ◽  
Zero Order Kinetics ◽  
Membrane Bound ◽  
Cyclic Phosphate

1. Breakdown of phosphatidylinositol was studied in homogenates and subcellular fractions of rat cerebral cortex by using both membrane-bound and externally added [32P]phosphatidylinositol as substrate. 2. In the presence of deoxycholate breakdown followed first-order kinetics at low substrate concentrations ([unk]1mm) and zero-order kinetics at higher concentrations (6–9mm). 3. Maximum breakdown by cerebral-cortex homogenates was approximately 0.5μmol/h per mg of protein and occurred at pH7.0 in the presence of 8mm-phosphatidylinositol, 2mm-CaCl2 and 2mg of deoxycholate/ml. Activity was abolished by 1mm-ethanedioxybis(ethylamine)tetra-acetate. 4. The products of phosphatidylinositol breakdown were 1,2-diacylglycerol and a mixture of d-myoinositol 1:2-cyclic phosphate (55%) and d-myoinositol 1-phosphate (45%). The two phosphate esters appeared to be produced together and in constant proportions. 5. Some 51% of the activity was particle-bound, with the highest activities in small nerve endings, microsomal material and two synaptic membrane fractions (fractions Mic20, Mic100, M1 1.0 and M1 0.9 respectively), all of which were also rich in acetylcholinesterase and which have been shown to be rich in other surface-membrane enzymes. Much of the particle-bound activity therefore appears to be present in cerebral-cortex plasma membranes. 6. The results are discussed in relation to previously described soluble activities that catalyse the same reaction, and to a possible role of the membrane-bound enzyme in enhanced phosphatidylinositol turnover in externally stimulated cells.

Role of lindane in membranes. Effects on membrane fluidity and activity of membrane-bound proteins

1994 ◽  
Vol 14 (3) ◽  
pp. 131-138 ◽  
Author(s):  
P. López-Aparicio ◽  
M. N. Recio ◽  
J. C. Prieto ◽  
M. A. Pérez-Albarsanz
Keyword(s):  
Membrane Fluidity ◽  
Adrenergic Receptor ◽  
Plasma Membranes ◽  
Opposite Effect ◽  
Glucose Transporter ◽  
Regulatory Mechanism ◽  
Cyclic Amp Accumulation ◽  
Membrane Bound ◽  
The Difference

The influence of lindane (gamma-hexachlorocyclohexane) on fluidity of plasma membranes from rat renal cortical tubules has been investigated. Preincubation with lindane increased membrane fluidity. This effect was accompanied by (i) a decrease in the transport of glucose with regard to the controls and (ii) an inhibition of the β-adrenergic stimulatory activity upon cyclic AMP accumulation. However, a significant decrease of the membrane fluidity was found when rats were injected with lindane for 12 days. The injection of lindane exerted the opposite effect on the membrane proteins, the glucose transporter and the β-adrenergic receptor, enhancing the glucose uptake and increasing the isoproterenol-stimulated cycle AMP accumulation. A possible explanation of the difference could involve a resistance to membrane disordering by lindane through a regulatory mechanism that would balance the activity of many lindane-sensitive proteins in insecticide-injected rats.

scholarly journals Modulation of lipid fluidity of small- and large-intestinal antipodal membranes by Ca2+

1986 ◽  
Vol 239 (3) ◽  
pp. 625-631 ◽  
Author(s):  
T A Brasitus ◽  
P K Dudeja
Keyword(s):  
Lipid Bilayers ◽  
Brush Border ◽  
Plasma Membranes ◽  
Bivalent Cation ◽  
Distal Small Intestine ◽  
Membrane Bound ◽  
Series Of Experiments ◽  
Membrane Bound Enzymes ◽  
Complex Manner

A series of experiments were conducted to examine the role of Ca2+ in modulating the fluidity of rat small- and large-intestinal antipodal plasma membranes and their liposomes. This bivalent cation was found to decrease the fluidity of these preparations in a complex manner involving at least two distinct mechanisms. The first appeared to be a direct effect of Ca2+ on fluidity, was readily reversible by addition of EGTA and presumably involved binding of Ca2+ to anionic sites in the lipid bilayers of these membranes. This effect was seen with all preparations examined. In contrast, the second effect of Ca2+ on fluidity was only seen in intact small-intestinal brush-border membranes, appeared to be indirect, was time- and cation-dependent, was only minimally reversible by addition of EGTA, and appeared to involve stimulation of membrane-bound enzymes which altered this membrane's fatty acid composition. Furthermore, regional differences in this latter effect of Ca2+ on brush-border membrane fluidity were also seen in these studies, i.e. proximal greater than distal small intestine.

scholarly journals Measurement of free and membrane-bound cathepsin G in human neutrophils using new sensitive fluorogenic substrates

2002 ◽  
Vol 366 (3) ◽  
pp. 965-970 ◽  
Author(s):  
Sylvie ATTUCCI ◽  
Brice KORKMAZ ◽  
Luiz JULIANO ◽  
Eric HAZOUARD ◽  
Catherine GIRARDIN ◽  
...  
Keyword(s):  
Cathepsin G ◽  
Polymorphonuclear Neutrophils ◽  
Human Neutrophils ◽  
Proteinase 3 ◽  
Protein Targets ◽  
Peptide Substrates ◽  
Loop Sequence ◽  
Membrane Bound

Activated human polymorphonuclear neutrophils at inflammatory sites release the chymotrypsin-like protease cathepsin G, together with elastase and proteinase 3 (myeloblastin), from their azurophil granules. The low activity of cathepsin G on synthetic substrates seriously impairs studies designed to clarify its role in tissue inflammation. We have solved this problem by producing new peptide substrates with intramolecularly quenched fluorescence. These substrates were deduced from the sequence of putative protein targets of cathepsin G, including the reactive loop sequence of serpin inhibitors and the N-terminal domain of the protease-activated receptor of thrombin, PAR-1. Two substrates were selected, Abz-TPFSGQ-EDDnp and Abz-EPFWEDQ-EDDnp, that are cleaved very efficiently by cathepsin G but not by neutrophil elastase or proteinase 3, with specificity constants (kcat/Km) in the 105M-1·s-1 range. They can be used to measure subnanomolar concentrations of free enzyme in vitro and at the surface of neutrophils purified from fresh human blood. Purified neutrophils express 0.02—0.7pg of cathepsin G/cell (n = 15) at their surface. This means that about 104 purified cells may be enough to record cathepsin G activity within minutes. This may be most important for investigating the role of cathepsin G as an inflammatory agent, especially in bronchoalveolar lavage fluids from patients with pulmonary inflammatory disorders.

Pathogenic and Compensatory Mechanisms in Epidermis of Sphingomyelin Synthase 2-Deficient Mice

2021 ◽  
pp. 1-7
Author(s):  
Shota Sakai ◽  
Asami Makino ◽  
Akihito Nishi ◽  
Takeshi Ichikawa ◽  
Tadashi Yamashita ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Lamellar Body ◽  
Lipid Mediator ◽  
Permeability Barrier ◽  
Compensatory Mechanisms ◽  
Terrestrial Environment ◽  
Sphingomyelin Synthase ◽  
Epidermal Permeability Barrier ◽  
Deficient Mice

Sphingomyelin (SM) is a constituent of cellular membranes, while ceramides (Cer) produced from SM on plasma membranes serve as a lipid mediator that regulates cell proliferation, differentiation, and apoptosis. In the skin, SM also is a precursor of Cer, an important constituent of epidermal permeability barrier. We investigated the role of epidermal SM synthase (SMS)2, an isoform of SMS, which modulates SM and Cer levels on plasma membranes. Although SMS2-knockout (SMS2-KO) mice were not neonatal lethal, an ichthyotic phenotype with epidermal hyperplasia and hyperkeratosis was evident at birth, which persisted until 2 weeks of age. These mice showed abnormal lamellar body morphology and secretion, and abnormal extracellular lamellar membranes in the stratum corneum. These abnormalities were no longer evident by 4 weeks of age in SMS2-KO mice. Our study suggests that (1) exposure to a dry terrestrial environment initiates compensatory responses, thereby normalizing epidermal ichthyotic abnormalities and (2) that a nonlethal gene abnormality can cause an ichthyotic skin phenotype.

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