Plasma Membrane Of Red Blood Cells Research Articles

The New Permeability Pathways: Targets and Selective Routes for the Development of New Antimalarial Agents

The malaria parasite, Plasmodium falciparum, spends part of its complex life cycle within the red blood cells of a human host. During this time, the parasite alters the permeability of the red blood cell's plasma membrane to allow the uptake of nutrients, the removal of "waste" and volume and ion regulation of the infected cell. The increased permeability is due to the induction of new permeability pathways (NPP), which are obvious chemotherapeutic antimalarial targets and/or selective routes for drugs, which target the internal parasite. This review covers our present understanding of the NPP, the methods used to screen for putative inhibitors of the NPP, the current repertoire of NPP inhibitors and the problems that need to be addressed to realise the potential of the NPP as antimalarial targets. In addition, the review will cover the use of the NPP as specific drug delivery routes.

Proteomic Analysis Identifies Novel Proteins of the Maurer's Clefts, a Secretory Compartment Delivering Plasmodium falciparum Proteins to the Surface of Its Host Cell

A novel method was validated for the efficient distinction between malaria parasite-derived and host cell proteins in mass spectrometry analyses. This method was applied to a ghost fraction from Plasmodium falciparum-infected erythrocytes containing the red blood cell plasma membrane, the erythrocyte submembrane skeleton, and the Maurer's clefts, a Golgi-like apparatus linked to and addressing parasite proteins to the host cell surface. This method allowed the identification of 78 parasite proteins. Among these we identified seven novel proteins of the Maurer's clefts based on immunofluorescence studies and proteinase K digestion assays. The products of six contiguous genes located on chromosome 5 were identified, and the location within the Maurer's clefts was established for two of them. This suggests a clustering of genes encoding Maurer's cleft proteins. Our study sheds new light on the biological function of the Maurer's clefts, which are central to the pathogenesis and to the intraerythrocytic development of P. falciparum.

New Technology for Red Blood Cell Preservation.

A method for freeze-drying red blood cells (RBCs) while maintaining a high degree of viability, has important implications in blood transfusion and clinical medicine. The disaccharide trehalose, found in animals capable of surviving dehydration can aid in this process. We are reporting a method for loading RBCs with trehalose followed by subsequent freeze-drying and rehydration. The loading of erythrocytes is based on the thermal properties of the RBC plasma membranes and provides efficient uptake of the sugar at 37°C in a time span of 7 hours. The data show that RBCs can be loaded with trehalose from the extracellular medium through a combination of osmotic imbalance and the phospholipid phase transition, producing an intracellular trehalose concentration of about 40 mM. Freeze-drying of trehalose loaded RBCs results in water contents in the range between 2 and 4 % and a level of survival of around 37 %, as measured by the extent of hemolysis. Surprisingly, freeze-dried and rehydrated RBCs showed high levels of ATP and 2,3-DPG and low methemoglobin. Biochemical analysis demonstrated that the activities of superoxide dismutase, catalase and acetylcholine esterase in freeze-dried RBCs are similar to those in fresh RBCs. These data provide an important step toward a stable erythrocyte product, which will be invaluable for transfusion and clinical applications. Supported by DARPA grant N66001-03-1-9827

Loading red blood cells with trehalose: a step towards biostabilization

A method for freeze-drying red blood cells (RBCs) while maintaining a high degree of viability has important implications in blood transfusion and clinical medicine. The disaccharide trehalose, found in animals capable of surviving dehydration can aid in this process. As a first step toward RBC preservation, we present a method for loading RBCs with trehalose. The method is based on the thermal properties of the RBC plasma membranes and provides efficient uptake of the sugar at 37 °C in a time span of 7 h. The data show that RBCs can be loaded with trehalose from the extracellular medium through a combination of osmotic imbalance and the phospholipid phase transition, resulting in intracellular trehalose concentrations of about 40 mM. During the loading period, the levels of ATP and 2,3-DPG are maintained close to the levels of fresh RBCs. Increasing the membrane fluidity through the use of a benzyl alcohol results in a higher concentration of intracellular trehalose, suggesting the importance of the membrane physical state for the uptake of the sugar. Osmotic fragility data show that trehalose exerts osmotic protection on RBCs. Flow cytometry data demonstrate that incubation of RBCs in a hypertonic trehalose solution results in a fraction of cells with different complexity and that it can be removed by washing and resuspending the RBCs in an iso-osmotic medium. The data provide an important first step in long-term preservation of RBCs.

Xenorhabdus nematophila (Enterobacteriacea) Secretes a Cation-selective Calcium-independent Porin Which Causes Vacuolation of the Rough Endoplasmic Reticulum and Cell Lysis

Xenorhabdus nematophila and Photorhabdus luminescens are two related enterobacteriaceae studied for their use in biological control and for synthesis of original virulence factors and new kinds of antibiotics. X. nematophila broth growth exhibits different cytotoxic activities on insect (Spodoptera littoralis, lepidoptera) immunocytes (hemocytes). Here we report the purification of the flhDC-dependent cytotoxin, a 10,790-Da peptide we have called alpha-Xenorhabdolysin (alpha X). We show that plasma membrane of insect hemocytes and of mammal red blood cells is the first target of this toxin. Electrophysiological and pharmacological approaches indicate that the initial effect of alpha X on macrophage plasma membrane is an increase of monovalent cation permeability, sensitive to potassium channel blockers. As a consequence, several events can occur intracellularly, such as selective vacuolation of the endoplasmic reticulum, cell swelling, and cell death by colloid-osmotic lysis. These effects, inhibited by potassium channel blockers, are totally independent of Ca(2+). However, the size of the pores created by alpha X on macrophage or red blood cell plasma membrane increases with toxin concentration, which leads to a rapid cell lysis.

Lipopolysaccharide from Proteus mirabilis O29 induces changes in red blood cell membrane lipids and proteins

Alterations in red blood cell (RBC) plasma membranes, i.e. in lipids and proteins, and osmotic fragility of these cells after treatment with Proteus mirabilis O29 endotoxin (lipolysaccharide (LPS)) were examined using a spin labelling method. At the highest concentration of LPS, insignificantly decreased fluidity of membrane lipids was observed. Changes in conformation of membrane proteins were determined by two covalently bound spin labels, 4-maleimido-2,2,6,6-tetramethylpiperidine-1-oxyl (MSL) and 4-iodoacetamido-2,2,6,6-tetramethylpiperidine-1-oxyl (ISL). The analysis of spectra of MSL and ISL showed modifications in membrane proteins in red blood cells treated with the highest concentration of lipopolysaccharide. On the other hand, in the case of isolated membranes, disturbances in membrane were observed for all concentrations of LPS. The alterations in membrane lipids and proteins are paralleled in a significant rise in osmotic fragility of RBCs upon endotoxin treatment. These results provide experimental evidence that P. mirabilis O29 LPS causes deleterious changes in membranes of human red blood cells. They show that action of lipopolysaccharide mainly concerns the membrane cytoskeleton.

Affinity driven molecular transfer from erythrocyte membrane to target cells

A wide variety of antimicrobial peptides are known to bind to - and disrupt microbial plasma membranes. Recently, derivatives of the antimicrobial peptide dermaseptin S4 were shown to selectively disrupt the plasma membrane of the intracellular parasite Plasmodium falciparum without harming that of the mammalian host cell. The resulting antimalarial activity is allegedly exerted after the harmless peptide binding to the membrane of the host cell, followed by peptide translocation across a number of intracellular membrane systems and interaction with that of the intraerythrocyte parasite. In this study, we present evidence in support of the ability of a membrane-bound peptide, the dermaseptin S4 derivative K 4-S4(1–13)a, to transfer from red blood cells (RBCs) to another distant membrane. Binding of K 4-S4(1–13)a to the plasma membrane of RBCs was assessed in vitro and in vivo, and found to be rapid, spontaneous and receptor independent, as was the transfer of the RBC-bound peptide to the plasma membrane of microorganisms. The present study further provides a basis for the possible use of RBCs as a transport vehicle to deliver drugs to distant targets. This drug delivery system involves the transient “loading” of RBCs with a lipophilic “hook” peptide. Such a peptide has enough affinity for the RBC’s plasma membrane to bind to the membrane, but given the opportunity, the peptide will exit its position and transfer to another (target) cell for which it has a greater affinity. The efficacy of such an affinity driven transfer system was demonstrated experimentally by the transfer of K 4-S4(1–13)a from pre-loaded RBCs to bacteria, yeast and protozoan target cells.

Association of Stomatin (Band 7.2b) with Glut1 Glucose Transporter

Employing a monoclonal antibody directed against the C-terminal peptide of glucose transporter molecule 1 (Glut1), we identified a ∼30-kDa polypeptide which coimmunoprecipitated with Glut1 from sample of human red blood cells (RBC) membranes. The ∼30-kDa polypeptide reacted with an antibody directed against stomatin, an integral plasma membrane protein which is also present at a high abundance in the human RBC plasma membrane. Likewise, employing anti-stomatin antibody, we found that Glut1 coimmunoprecipitated with stomatin from samples of RBC membranes. However, neither band 3, which is the most abundant integral membrane protein in the RBC, nor actin coimmunoprecipitated with Glut1, indicating a specific interaction between Glut1 and stomatin. Similar to the results obtained in the RBC, Glut1 and stomatin immunoprecipitated with each other in lysates of Clone 9 cells, a rat liver cell line in which Glut1 is expressed at ∼1/200 the level present in RBC. Employing conditions that resulted in immunoprecipitation of ∼10% of Glut1 in RBC membranes led to a ∼3% coimmunoprecipitation of stomatin. A mixed population of Clone 9 cells stably transfected with a plasmid overexpressing the mouse stomatin exhibited 30 ± 3% reduction in the basal rate of glucose transport compared to control cells or cells stably transfected with the empty vector. The above results suggest that stomatin is closely associated with Glut1 in the plasma membrane and that overexpression of stomatin results in a depression in the basal rate of glucose transport.

Effects of salicylic acid derivatives on red blood cell membranes.

Salicylamide, sodium salicylate and acetylsalicylic acid are salicylic acid derivates. They differ in their substitution on the benzene ring and may have different effects on membranes. Red blood cells were used as a prototypical cellular system regarding drug mediated plasma bilayer effects. Established photometric methods sensing tiny changes of red blood cell morphology at rest (red blood cell shape) and at very low shear forces (red blood cell stiffness, red blood cell relaxation time) were applied. The derivative induced effects were detected in a time- and dose-dependent manner. Salicylamide induced a most pronounced echinocytic shape at 5 mM. The shape effect was smaller above as well as below 5 mM. Sodium salicylate induced echinocytes with increasing concentrations showing a saturation above 10 mM. In contrast, the shape was not affected by acetylsalicylic acid. All shape changes occurred within 2 min, and were reversible. The above tendencies were in parallel to a slight red blood cell stiffening. The relaxation time continuously increased with increasing concentrations in both salicylamide and sodium salicylate, with salicylamide always acting stronger. Acetylsalicylic acid again showed no effect. We hypothesize that the observed effects of sodium salicylate and salicylamide are due to their phenolic character mediating a molecular hydrophobicity. According to the bilayer couple hypothesis this would lead to an insertion into the red blood cells outer plasma bilayer leaflet. The extension induced here would cause a positive membrane bending leading to echinocytic shapes and the observed loss of red blood cell fluidity. In contrast, the hydrophilic aspirin would penetrate and thus not affect the red blood cell plasma membrane.

Effect of zinc on carp ( Cyprinus carpio L.) erythrocytes

The effect of zinc exposure on some properties of the carp erythrocyte membrane was studied in vitro. Red blood cells plasma membranes were separated from other cellular membranes using a combination of differential and density gradient centrifugation. The purity of obtained plasma membrane preparations was determined by measuring the activity of the marker enzymes. Electrophoretic patterns of the main erythrocyte membrane proteins excluded their degradation during the isolation and purification procedure. Carp erythrocyte membranes, obtained from cells previously incubated with increasing ZnSO 4 concentrations, were used to elucidate the effect of zinc ions on their physical and biochemical properties. Using fluorescent probes: 12-AS and TMA–DPH, we found that zinc ions reduced the fluidity of the lipid bilayer, both in the middle and near the aqueous interface. Moreover, it was observed that zinc had no significant influence neither on the Na,K-ATPase activity nor on the thiol groups content in the erythrocyte membrane. We also detected that incubation of erythrocytes with zinc lead to the marked decrease of hemolytic resistance of the cells. Our studies demonstrate that zinc at higher concentrations may be toxic to carp erythrocytes causing changes in the membrane fluidity and hemolytic resistance.

Kinetic models and phenomenological analysis of passive lipid translocation in single-file.

Passive movement of lipids through a membrane-embedded pore is analysed with kinetic equations of transport in single-file. The number of lipids arranged along the translocation coordinate in the pore is not limited in the calculations. The assumption is made that the energetic state of a pore is independent of the sequence of lipids contained in it. The results are valid for an arbitrary number of species with identical kinetic constants. It is shown that infinitely fast diffusion of one vacant site is equivalent to the filled pore approximation, which has been used here. We introduce the concept of non-strict single-file, which allows also for exchanges of neighbouring lipids inside the pore at specified rates. The model successfully simulates the redistribution of lipids between the monolayers of red blood cell plasma membranes under operation of an active aminophospholipid translocase. Kinetic equations are related to linear flux force relations. Phenomenological coefficients are expressed and analysed in terms of kinetic constants. Plausible kinetic pore model parameters are derived from comparison with a reference simulation of a thermodynamic model of the erythrocyte transmembrane lipid distribution. Mechanical forces due to differences in compressions of the lipid molecules between the monolayers are incorporated in kinetic rate constants. It is seen how the active inward transport of aminophospholipids causes an unsymmetrical passive redistribution of the other components due to mechanical effects and cross-coupling of fluxes.

Effects of urea and trimethylamine N-oxide on fluidity of liposomes and membranes of an elasmobranch.

The effects on membrane fluidity of two solutes of biological importance in elasmobranch fishes, urea and trimethylamine oxide (TMAO), were determined using elasmobranch red blood cell plasma membranes and artificial liposomes. Fluorescence polarizations of three probes with differing sites of insertion (1, 6-diphenylhexatriene, cis-parinaric acid, and trans-parinaric acid) were used to study the effects of physiological levels of urea (400 mM) and TMAO (200 mM) separately and together in a 2:1 urea:TMAO ratio (400 mM:200 mM). In the elasmobranch erythrocyte membrane, there was a trend toward an increase in the order of the gel-phase domains when treated with urea, although this was not statistically significant. This effect was counteracted by the presence of TMAO. To determine if the organic solutes were acting directly on the membrane lipids or on the integral proteins, phase-transition profiles of protein-free dipalmitoyl phosphatidylcholine liposomes were determined. These profiles showed that urea again increased the order of the gel-phase domains of the bilayer; however, this effect was not counteracted by the presence of TMAO. We suggest that the increased order in the gel-phase domains may be an indirect effect of a decrease in the order of the fluid-phase domains. This increase in fluidity may be due either to a disruptive effect of urea on the hydrophobic core of the membrane or to indirect effects mediated by changes in the integral membrane proteins. This study is the first to demonstrate that urea and TMAO may act as counteracting solutes in the elasmobranch erythrocyte membrane and that the counteraction appears to be at the level of the integral proteins rather than the membrane lipids.

Activation of Glut1 Glucose Transporter in Human Erythrocytes

Glut1, the only glucose transporter isoform expressed in the human red blood cell (RBC), binds to and is inhibited by cytochalasin B (CB). In the present study we show that incubation of RBC ghost membranes with 10 μM cytochalasin E (CE) results in a 1.8-fold increase in the number of glucose-sensitive cytochalasin B binding sites. Moreover, treatment with CE was associated with no observable change in the protein composition of RBC ghosts determined by SDS–PAGE. Removal of surface (“extrinsic”) proteins from RBC ghosts by treatment with 0.2 mM EDTA (pH 12) for 30 min resulted in a similar 1.8-fold increase in the number of glucose-sensitive CB binding sites. Western blot analysis showed that treatment with CE or EDTA resulted in no change in the amount or mobility of Glut1 present in ghost membranes. Glucose transport, measured as CB-inhibitable 3-O-[3H]methylglucose uptake by resealed ghosts, was stimulated in CE-treated resealed ghosts, with thet1/2to equilibrium decreasing from 6.8 ± 0.5 to 3.9 ± 0.3 s (P< 0.05). Treatment of ghosts with CE or EDTA followed by Western blotting of samples in the presence or absence of β-mercaptoethanol resulted in no change in immunoreactivity or mobility of the major Glut1 band. The above results suggest that a significant fraction of Glut1 transporters exists in an inactive form (“masked”) in RBC plasma membranes and that treatment of ghosts with CE or EDTA leads to an apparent activation of Glut1.

Amphiphile induced echinocyte-spheroechinocyte transformation of red blood cell shape.

A possible physical explanation of the echinocyte -spheroechinocyte red blood cell (RBC) shape transformation induced by the intercalation of amphiphilic molecules into the outer layer of the RBC plasma membrane bilayer is given. The stable RBC shape is determined by the minimization of the membrane elastic energy, consisting of the bilayer bending energy, the bilayer relative stretching energy and the skeleton shear elastic energy. It is shown that for a given relative cell volume the calculated number of echinocyte spicula increases while their size decreases as the number of the intercalated amphiphilic molecules in the outer layer of the cell membrane bilayer is increased, which is in agreement with experimental observations. Further, it is show that the equilibrium difference between the outer and the inner membrane leaflet areas of the stable RBC shapes increases if the amount of the intercalated amphiphiles is increased, thereby verifying theoretically the original bilayer couple hypothesis of Sheetz and Singer (1974) and Evans (1974).

Complete Deficiency of Glycophorin A in Red Blood Cells From Mice With Targeted Inactivation of the Band 3 (AE1) Gene

AbstractGlycophorin A is the major transmembrane sialoglycoprotein of red blood cells. It has been shown to contribute to the expression of the MN and Wright blood group antigens, to act as a receptor for the malaria parasite Plasmodium falciparum and Sendai virus, and along with the anion transporter, band 3, may contribute to the mechanical properties of the red blood cell membrane. Several lines of evidence suggest a close interaction between glycophorin A and band 3 during their biosynthesis. Recently, we have generated mice where the band 3 expression was completely eliminated by selective inactivation of the AE1 anion exchanger gene, thus allowing us to study the effect of band 3 on the expression of red blood cell membrane proteins. In this report, we show that the band 3 −/− red blood cells contain protein 4.1, adducin, dematin, p55, and glycophorin C. In contrast, the band 3 −/− red blood cells are completely devoid of glycophorin A (GPA), as assessed by Western blot and immunocytochemistry techniques, whereas the polymerase chain reaction (PCR) confirmed the presence of GPA mRNA. Pulse-label and pulse-chase experiments show that GPA is not incorporated in the membrane and is rapidly degraded in the cytoplasm. Based on these findings and other published evidence, we propose that band 3 plays a chaperone-like role, which is necessary for the recruitment of GPA to the red blood cell plasma membrane.

Complete Deficiency of Glycophorin A in Red Blood Cells From Mice With Targeted Inactivation of the Band 3 (AE1) Gene

Glycophorin A is the major transmembrane sialoglycoprotein of red blood cells. It has been shown to contribute to the expression of the MN and Wright blood group antigens, to act as a receptor for the malaria parasite Plasmodium falciparum and Sendai virus, and along with the anion transporter, band 3, may contribute to the mechanical properties of the red blood cell membrane. Several lines of evidence suggest a close interaction between glycophorin A and band 3 during their biosynthesis. Recently, we have generated mice where the band 3 expression was completely eliminated by selective inactivation of the AE1 anion exchanger gene, thus allowing us to study the effect of band 3 on the expression of red blood cell membrane proteins. In this report, we show that the band 3 −/− red blood cells contain protein 4.1, adducin, dematin, p55, and glycophorin C. In contrast, the band 3 −/− red blood cells are completely devoid of glycophorin A (GPA), as assessed by Western blot and immunocytochemistry techniques, whereas the polymerase chain reaction (PCR) confirmed the presence of GPA mRNA. Pulse-label and pulse-chase experiments show that GPA is not incorporated in the membrane and is rapidly degraded in the cytoplasm. Based on these findings and other published evidence, we propose that band 3 plays a chaperone-like role, which is necessary for the recruitment of GPA to the red blood cell plasma membrane.

Transport and trafficking in malaria-infected erythrocytes

D espite almost a century of effort aimed at eradication, malaria remains one of the world's major health problems. The disease is caused by the eukaryotic apicomplexan parasite Plasrnodium spp. The most virulent of the four species known to infect humans is Plasmodium falciparum, and we will focus on this species alone. The pathology of the disease results from the multiplication of asexual parasites in red blood cells (RBCs) and the sequestration of these infected cells in the organs of the vertebrate host. The metabolic processes leading to parasite maturation at this stage of the life cycle are poorly understood. There have been many recent studies on the mechanisms of uptake of metabolites in vitro by asexual parasites, the most recent of which is by Haldar's group I and attempts to characterize an uptake route that could constitute a target for either chemotherapy or drug delivery. Asexual stages of the parasite mature in RBCs within a parasitophorous vacuole (PV) formed during merozoite invasion. It has long been recognized that the parasite imports host cell haemoglobin in a macropinocytotic process involving an invagination of the parasite plasmalemma with host cell cytoplasm at a specialized structure termed the micropore 2 (Fig. 1). However, not all nutrients and growth factors necessary for parasite maturation are provided by the digestion of haemoglobin: in addition, several exogenous substrates, including glucose, assorted dicarboxylic acids, purines and specific amino acids 3,4 that are essential for parasite maturation, are imported from the extracellular milieu 3,s. Thus, although the host RBC provides a protected environment, the intracellular location of the parasite requires imported and exported solutes to cross a series of three unit membranes: the RBC plasmalemma, the PV membrane and the parasite plasmalemma. The mechanisms by which malaria parasites obtain nutrients have been the subject of much controversy and debate during the past few years. There have been reports of three potential mechanisms for uptake: (1) by parasite-induced permeability changes in the RBC plasmalemma (for reviews, see Refs 3,6); (2) by a direct connection known as a parasitophorous duct between the parasite, and the external milieu7,8; and (3) by the involvement of a tubovesicular membrane (TVM) network in directed transport across the RBC cytoplasm *,3,9. Parasiteinduced permeability changes in RBC plasma membranes, which occur 12-20 h postinvasion, have been described by several groups (reviewed in Refs 3,6). The existence and possible role of the parasitophorous duct is still controversial and lacks independent corroboration. Indeed, various ukrastructural studies using electron-dense tracers and labelling techniques ~°,~1 have so far failed to find any evidence for a direct connection between the exterior of the RBC and any of the intracellular compartments. Even the most recent paper by the Taraschi group 12, which claims to provide ultrastructural evidence for the presence of the duct, does not actually give illustrations showing membrane continuity between the TVIVl and the RBC plasmalemma (as the

Oxidative injury and potential use of antioxidants in schizophrenia

There is increasing evidence that oxidative injury contributes to pathophysiology of schizophrenia, indicated by the increased lipid peroxidation products in plasma and CSF, and altered levels of both enzymatic and non-enzymatic antioxidants in chronic and drug-naive first-episode schizophrenic patients. The increased plasma lipid peroxidation is also supported by concomitant lower levels of esterified polyunsaturated essential fatty acids of red blood cell plasma membrane phospholipids. Because membrane phospholipids play a critical role in neuronal signal transduction, oxidative damage of these lipids may contribute to the proposed altered neurotransmitter receptor-mediated signal transduction and thereby alter information processing in schizophrenia. Adjunctive treatment with antioxidants (e.g. vitamins E and C, β-carotene and quinones) at the initial stages of illness may prevent further oxidative injury and thereby ameliorate and prevent further possible deterioration of associated neurological and behavioral deficits in schizophrenia.

Mechanism of inhibition of the plasma membrane Ca(2+)-ATPase by barbiturates.

We have demonstrated that sodium pentobarbital inhibited the activation of the human red blood cell plasma membrane Ca(2+)-ATPase produced by dimerization of enzyme monomers or by calmodulin binding to enzyme monomers. The effects of the barbiturate were dose-dependent. Both Vmax and Ca2+ affinity were reduced. The Ca(2+)-ATPase activity of the dimeric enzyme was distinctly less sensitive with respect to the effective inhibitory concentrations of pentobarbital and to the rate of onset of inhibition than was the calmodulin-dependent activation of enzyme monomers. Temperature dependence of the inhibition was in agreement with direct, nonpolar interactions of pentobarbital with a water-exposed nonpolar patch on the surface of this transmembrane protein. The barbiturate prevented the increase of intrinsic tryptophan fluorescence associated with substrate Ca2+ binding to the enzyme dimer. On the basis of the barbiturate effects we propose a model for the action of detergent-like compounds on the enzyme. They inhibit Ca(2+)-ATPase activity by binding to a nonpolar patch on the water-exposed dimerization surface of the enzyme monomer, part of which is also the binding site for calmodulin. The model assumes that their binding to the nonpolar patch on the monomer interferes with dimerization and weakens but does not prohibit calmodulin binding, whose activation of the enzyme is then submaximal. The model should be applicable to other proteins as the two activation pathways studied have been demonstrated for various enzymes.

Plasmodium falciparum: Highly Mobile Small Vesicles in the Malaria-Infected Red Blood Cell Cytoplasm

A dynamic population of small vesicles within the cytoplasm of live malaria-infected red blood cells has been studied using a laser scanning confocal microscope. Acridine orange was used to follow the movement of vesicles within the infected red blood cell cytoplasm, including the budding of vesicles from the malarial parasite. These highly mobile vesicles are found predominantly in mid- to late ring-stage parasites and are almost entirely absent from young rings and mature trophozoites. Since the known parasite modifications of the red blood cell plasma membrane in mid-ring- to early trophozoite-stage parasites correlates with the appearance of acridine orange-staining vesicles, these vesicles may be an important component in the transport of parasite proteins across the infected red blood cell cytoplasm.