The Experts below are selected from a list of 165 Experts worldwide ranked by ideXlab platform
Samuel E Lux - One of the best experts on this subject based on the ideXlab platform.
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Protein 4.2 binds to the carboxyl terminal ef hands of erythroid α spectrin in a calcium and calmodulin dependent manner
Journal of Biological Chemistry, 2010Co-Authors: Catherine Korsgren, Luanne L. Peters, Samuel E LuxAbstract:Spectrin and Protein 4.1 cross-link F-actin protofilaments into a network called the membrane skeleton. Actin and 4.1 bind to one end of β-spectrin. The adjacent end of α-spectrin, called the EF-domain, is calmodulin-like, with calcium-dependent and calcium-independent EF-hands. It has no known function. However, the sph1J/sph1J mouse has very fragile red cells and lacks the last 13 amino acids in the EF-domain, suggesting the domain is critical for skeletal integrity. Using pulldown binding assays, we find the α-spectrin EF-domain either alone or incorporated into a mini-spectrin binds native and recombinant Protein 4.2 at a previously identified region of 4.2 (G3 peptide). Native 4.2 binds with an affinity comparable with other membrane skeletal interactions (Kd = 0.30 μm). EF-domains bearing the sph1J mutation are inactive. Binding of Protein 4.2 to band 3 (Kd = 0.45 μm) does not interfere with the spectrin-4.2 interaction. Spectrin-4.2 binding is amplified by micromolar concentrations of Ca2+ (but not Mg2+) by three to five times. Calmodulin also binds to the EF-domain (Kd = 17 μm), and Ca2+-calmodulin blocks Ca2+-dependent binding of Protein 4.2 but not Ca2+-independent binding. The data suggest that Protein 4.2 is located near Protein 4.1 at the spectrin-actin junctions. Because Proteins 4.1 and 4.2 also bind to band 3, the erythrocyte anion channel, we suggest that one or both of these Proteins cause a portion of band 3 to localize near the spectrin-actin junctions and provide another point of attachment between the membrane skeleton and the lipid bilayer.
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hereditary spherocytosis defects in Proteins that connect the membrane skeleton to the lipid bilayer
Seminars in Hematology, 2004Co-Authors: Stefan W Eber, Samuel E LuxAbstract:The molecular causes of hereditary spherocytosis (HS) have been unraveled in the past decade. No frequent defect is found, and nearly every family has a unique mutation. In dominant HS, nonsense and frameshift mutations of ankyrin, band 3, and beta-spectrin predominate. Recessive HS is most often due to compound heterozygosity of defects in ankyrin, alpha-spectrin, or Protein 4.2. Common combinations include a defect in the promoter or 5'-untranslated region of ankyrin paired with a missense mutation, a low expression allele of alpha-spectrin plus a missense mutation, and various mutations in the gene for Protein 4.2. In most patients' red cells, no abnormal Protein is present. Only rare missense mutations, like ankyrin Walsrode (V463I) or beta-spectrin Kissimmee (W202R), have given any insight into the functional domains of the respective Proteins. Although the eminent role of the spleen in the premature hemolysis of red cells in HS is unquestioned, the molecular events that cause splenic conditioning of spherocytes are unclear. Electron micrographs show that small membrane vesicles are shed during the formation of spherocytes. Animal models give further insight into the pathogenetic consequences of membrane Protein defects as well as the causes of the variability of disease severity.
Stefan W Eber - One of the best experts on this subject based on the ideXlab platform.
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hereditary spherocytosis defects in Proteins that connect the membrane skeleton to the lipid bilayer
Seminars in Hematology, 2004Co-Authors: Stefan W Eber, Samuel E LuxAbstract:The molecular causes of hereditary spherocytosis (HS) have been unraveled in the past decade. No frequent defect is found, and nearly every family has a unique mutation. In dominant HS, nonsense and frameshift mutations of ankyrin, band 3, and beta-spectrin predominate. Recessive HS is most often due to compound heterozygosity of defects in ankyrin, alpha-spectrin, or Protein 4.2. Common combinations include a defect in the promoter or 5'-untranslated region of ankyrin paired with a missense mutation, a low expression allele of alpha-spectrin plus a missense mutation, and various mutations in the gene for Protein 4.2. In most patients' red cells, no abnormal Protein is present. Only rare missense mutations, like ankyrin Walsrode (V463I) or beta-spectrin Kissimmee (W202R), have given any insight into the functional domains of the respective Proteins. Although the eminent role of the spleen in the premature hemolysis of red cells in HS is unquestioned, the molecular events that cause splenic conditioning of spherocytes are unclear. Electron micrographs show that small membrane vesicles are shed during the formation of spherocytes. Animal models give further insight into the pathogenetic consequences of membrane Protein defects as well as the causes of the variability of disease severity.
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characteristic features of the genotype and phenotype of hereditary spherocytosis in the japanese population
International Journal of Hematology, 2000Co-Authors: Yoshihito Yawata, Akiko Kanzaki, Ayumi Yawata, Walter Doerfler, Refik Ozcan, Stefan W EberAbstract:Abstract Hereditary spherocytosis (HS) is the most common hemolytic anemia of congenital origin in the Japanese population. Among 844 cases of 520 kindred with congenital red cell membrane disorders studied at the Kawasaki Medical School in the last 25 years (1975-1999), 407 cases (48.2%) of 215 kindred had HS. Among the recent 60 kindred with HS, autosomal dominant (AD) transmission was proven in 19. The remaining 41 non-AD HS included 1) homozygous patients with autosomal recessive inheritance, 2) HS patients with de novo gene mutations, and 3) mild HS with AD inheritance. The extent of clinical severity in the non-AD HS cases was nearly identical to that in the AD cases. The incidence of membrane Protein abnormalities in our 60 Japanese HS kindred was unique: there were lower ankyrin deficiencies (7%), moderate band 3 deficiencies (20%), and much higher Protein 4.2 deficiencies (45%), with 28% of unknown etiology. The incidence of membrane Protein deficiencies corresponded to that determined by gene analyses; i.e., mutations mostly in band 3 and/or in Protein 4.2 genes and fewer ankyrin gene mutations. In the band 3 gene, 11 mutations pathognomonic for HS were identified (3 frameshift and 8 missense mutations). There were 5 mutations of the Protein 4.2 gene (3 missense mutations, 1 nonsense mutation, and 1 splicing mutation) pathognomonic for HS. On the other hand, 2 missense mutations were detected in the ankyrin gene in this study. The genetic abnormalities in our HS patients correlated well with the phenotypic ultrastructural abnormalities of red cell membranes in situ. Ankyrin mutations (ankyrin Marburg and ankyrin Stuttgart with frameshift mutations) were associated mostly with a disrupted cytoskeletal network, and band 3 mutations (band 3 Kagoshima with frameshift mutation) typically demonstrated anomalies of intramembrane particles (IMPs). Protein 4.2 mutations (homozygotes of Protein 4.2 Nippon) with complete Protein 4.2 deficiency showed abnormalities of both the cytoskeletal network and IMPs.
Philip S. Low - One of the best experts on this subject based on the ideXlab platform.
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identification of the components of a glycolytic enzyme metabolon on the human red blood cell membrane
Journal of Biological Chemistry, 2013Co-Authors: Estela Puchulucampanella, Haiyan Chu, David J Anstee, Jacob A Galan, Andy W Tao, Philip S. LowAbstract:Glycolytic enzymes (GEs) have been shown to exist in multienzyme complexes on the inner surface of the human erythrocyte membrane. Because no Protein other than band 3 has been found to interact with GEs, and because several GEs do not bind band 3, we decided to identify the additional membrane Proteins that serve as docking sites for GE on the membrane. For this purpose, a method known as “label transfer” that employs a photoactivatable trifunctional cross-linking reagent to deliver a biotin from a derivatized GE to its binding partner on the membrane was used. Mass spectrometry analysis of membrane Proteins that were biotinylated following rebinding and photoactivation of labeled GAPDH, aldolase, lactate dehydrogenase, and pyruvate kinase revealed not only the anticipated binding partner, band 3, but also the association of GEs with specific peptides in α- and β-spectrin, ankyrin, actin, p55, and Protein 4.2. More importantly, the labeled GEs were also found to transfer biotin to other GEs in the complex, demonstrating for the first time that GEs also associate with each other in their membrane complexes. Surprisingly, a new GE binding site was repeatedly identified near the junction of the membrane-spanning and cytoplasmic domains of band 3, and this binding site was confirmed by direct binding studies. These results not only identify new components of the membrane-associated GE complexes but also provide molecular details on the specific peptides that form the interfacial contacts within each interaction.
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membranes in wild type and membrane Protein knockout mice Characterization of glycolytic enzyme interactions with murine erythrocyte
2013Co-Authors: Philip S. Low, M. Estela Campanella, Haiyan Chu, Luanne L. Peters, Nancy J Wandersee, Narla MohandasAbstract:Abstract Previous research has shown that glycolytic enzymes (GEs) exist as multi-enzyme complexes on the inner surface of human erythrocyte membranes. Because GE binding sites have been mapped to sequences on the membrane Protein, band 3, that are not conserved in other mammalian homologs, the question arose whether GEs can organize into complexes on other mammalian erythrocyte membranes. To address this, murine erythrocytes were stained with antibodies to glyceraldehyde-3-phosphate dehydrogenase, aldolase, phosphofructokinase, lactate dehydrogenase and pyruvate kinase and analyzed by confocal microscopy. GEs were found to localize to the membrane in oxygenated erythrocytes, but redistributed to the cytoplasm upon deoxygenation, as seen in human erythrocytes. To identify membrane Proteins involved in GE assembly, erythrocytes from mice lacking each of the major erythrocyte membrane Proteins were examined for GE localization. GEs from band 3 knockout mice were not membrane associated, but distributed throughout the cytoplasm, regardless of erythrocyte oxygenation state. In contrast, erythrocytes from mice lacking -spectrin, ankyrin, Protein 4.2, Protein 4.1, -adducin or dematin headpiece exhibited GEs bound to the membrane. These data suggest that oxygenation-dependent assembly of GEs on the membrane could be a general phenomenon of mammalian erythrocytes and that stability of these interactions depends primarily on band 3.
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Characterization of glycolytic enzyme interactions with murine erythrocyte membranes in wild-type and membrane Protein knockout mice
Blood, 2008Co-Authors: M. Estela Campanella, Haiyan Chu, Diana M. Gilligan, Luanne L. Peters, Nancy J Wandersee, Narla Mohandas, Philip S. LowAbstract:Previous research has shown that glycolytic enzymes (GEs) exist as multienzyme complexes on the inner surface of human erythrocyte membranes. Because GE binding sites have been mapped to sequences on the membrane Protein, band 3, that are not conserved in other mammalian homologs, the question arose whether GEs can organize into complexes on other mammalian erythrocyte membranes. To address this, murine erythrocytes were stained with antibodies to glyceraldehyde-3-phosphate dehydrogenase, aldolase, phosphofructokinase, lactate dehydrogenase, and pyruvate kinase and analyzed by confocal microscopy. GEs were found to localize to the membrane in oxygenated erythrocytes but redistributed to the cytoplasm upon deoxygenation, as seen in human erythrocytes. To identify membrane Proteins involved in GE assembly, erythrocytes from mice lacking each of the major erythrocyte membrane Proteins were examined for GE localization. GEs from band 3 knockout mice were not membrane associated but distributed throughout the cytoplasm, regardless of erythrocyte oxygenation state. In contrast, erythrocytes from mice lacking α-spectrin, ankyrin, Protein 4.2, Protein 4.1, β-adducin, or dematin headpiece exhibited GEs bound to the membrane. These data suggest that oxygenation-dependent assembly of GEs on the membrane could be a general phenomenon of mammalian erythrocytes and that stability of these interactions depends primarily on band 3.
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crystallographic structure and functional interpretation of the cytoplasmic domain of erythrocyte membrane band 3
Blood, 2000Co-Authors: Dachuan Zhang, Anatoly Kiyatkin, Jeffrey T Bolin, Philip S. LowAbstract:The red blood cell membrane (RBCM) is a primary model for animal cell plasma membranes. One of its major organizing centers is the cytoplasmic domain of band 3 (cdb3), which links multiple Proteins to the membrane. Included among its peripheral Protein ligands are ankyrin (the major bridge to the spectrin-actin skeleton), Protein 4.1, Protein 4.2, aldolase, glyceraldehyde-3-phosphate dehydrogenase, phosphofructokinase, deoxyhemoglobin, p72syk Protein tyrosine kinase, and hemichromes. The crystal structure of cdb3 is reported at 0.26 nm (2.6 A) resolution. A tight symmetric dimer is formed by cdb3; it is stabilized by interlocked dimerization arms contributed by both monomers. Each subunit also includes a larger peripheral Protein binding domain with an α+ β-fold. The binding sites of several peripheral Proteins are localized in the structure, and the nature of the major conformational change that regulates membrane-skeletal interactions is evaluated. An improved structural definition of the Protein network at the inner surface of the RBCM is now possible.
Stefan Eber - One of the best experts on this subject based on the ideXlab platform.
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hereditary spherocytosis defects in Proteins that connect the membrane skeleton to the lipid bilayer
Seminars in Hematology, 2004Co-Authors: Stefan EberAbstract:Abstract The molecular causes of hereditary spherocytosis (HS) have been unraveled in the past decade. No frequent defect is found, and nearly every family has a unique mutation. In dominant HS, nonsense and frameshift mutations of ankyrin, band 3, and β-spectrin predominate. Recessive HS is most often due to compound heterozygosity of defects in ankyrin, α-spectrin, or Protein 4.2. Common combinations include a defect in the promoter or 5′-untranslated region of ankyrin paired with a missense mutation, a low expression allele of α-spectrin plus a missense mutation, and various mutations in the gene for Protein 4.2. In most patients’ red cells, no abnormal Protein is present. Only rare missense mutations, like ankyrin Walsrode (V463I) or β-spectrin Kissimmee (W202R), have given any insight into the functional domains of the respective Proteins. Although the eminent role of the spleen in the premature hemolysis of red cells in HS is unquestioned, the molecular events that cause splenic conditioning of spherocytes are unclear. Electron micrographs show that small membrane vesicles are shed during the formation of spherocytes. Animal models give further insight into the pathogenetic consequences of membrane Protein defects as well as the causes of the variability of disease severity.
Reinhart A F Reithmeier - One of the best experts on this subject based on the ideXlab platform.
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Protein 4.2 interaction with hereditary spherocytosis mutants of the cytoplasmic domain of human anion exchanger 1
Biochemical Journal, 2011Co-Authors: Susan P Bustos, Reinhart A F ReithmeierAbstract:AE1 (anion exchanger 1) and Protein 4.2 associate in a Protein complex bridging the erythrocyte membrane and cytoskeleton; disruption of the complex results in unstable erythrocytes and HS (hereditary spherocytosis). Three HS mutations (E40K, G130R and P327R) in cdAE1 (the cytoplasmic domain of AE1) occur with deficiencies of Protein 4.2. The interaction of wild-type AE1, AE1HS mutants, mdEA1 (the membrane domain of AE1), kAE1 (the kidney isoform of AE1) and AE1SAO (Southeast Asian ovalocytosis AE1) with Protein 4.2 was examined in transfected HEK (human embryonic kidney)-293 cells. The HS mutants had wild-type expression levels and plasma-membrane localization. Protein 4.2 expression was not dependent on AE1. Protein 4.2 was localized throughout the cytoplasm and co-localized at the plasma membrane with the HS mutants mdAE1 and kAE1, but at the ER (endoplasmic reticulum) with AE1SAO. Pull-down assays revealed diminished levels of Protein 4.2 associated with the HS mutants relative to AE1. The mdAE1 did not bind Protein 4.2, whereas kAE1 and AE1SAO bound wild-type amounts of Protein 4.2. A Protein 4.2 fatty acylation mutant, G2A/C173A, had decreased plasma-membrane localization compared with wild-type Protein 4.2, and co-expression with AE1 enhanced its plasma-membrane localization. Subcellular fractionation showed the majority of wild-type and G2A/C173A Protein 4.2 was associated with the cytoskeleton of HEK-293 cells. The present study shows that cytoplasmic HS mutants cause impaired binding of Protein 4.2 to AE1, leaving Protein 4.2 susceptible to loss during erythrocyte development.
