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J C Kader

Publications and source records attributed to J C Kader.

At least 37 records · Page 2Linked to original sources
PubMed1994

A phosphatidylinositol/phosphatidylcholine transfer protein is required for differentiation of the dimorphic yeast Yarrowia lipolytica from the yeast to the mycelial form.

The SEC14SC gene encodes the phosphatidylinositol/phosphatidylcholine transfer protein (PI/PC-TP) of Saccharomyces cerevisiae. The SEC14SC gene product (SEC14pSC) is associated with the Golgi complex as a peripheral membrane protein and plays an essential role in stimulating Golgi secretory function. We report the characterization of SEC14YL, the structural gene for the PI/PC-TP of the dimorphic yeast Yarrowia lipolytica. SEC14YL encodes a primary translation product (SEC14YL) that is predicted to be a 497-residue polypeptide of which the amino-terminal 300 residues are highly homologous to the entire SEC14pSC, and the carboxyl-terminal 197 residues define a dispensible domain that is not homologous to any known protein. In a manner analogous to the case for SEC14pSC, SEC14pYL localizes to punctate cytoplasmic structures in Y. lipolytica that likely represent Golgi bodies. However, SEC14pYL is neither required for the viability of Y. lipolytica nor is it required for secretory pathway function in this organism. This nonessentiality of SEC14pYL for growth and secretion is probably not the consequence of a second PI/PC-TP activity in Y. lipolytica as cell-free lysates prepared from delta sec14YL strains are devoid of measurable PI/PC-TP activity in vitro. Phenotypic analyses demonstrate that SEC14pYL dysfunction results in the inability of Y. lipolytica to undergo the characteristic dimorphic transition from the yeast to the mycelial form that typifies this species. Rather, delta sec14YL mutants form aberrant pseudomycelial structures as cells enter stationary growth phase. The collective data indicate a role for SEC14pYL in promoting the differentiation of Y. lipolytica cells from yeast to mycelia, and demonstrate that PI/PC-TP function is utilized in diverse ways by different organisms.

Amino Acid Sequence↗
PubMed1993

Four 9-kDa proteins excreted by somatic embryos of grapevine are isoforms of lipid-transfer proteins.

Four 9-kDa small extracellular proteins produced by embryogenic cultures in the absence of auxin have been purified from the extracellular medium of grapevine somatic embryo cultures through cation-exchange chromatography and hydrophobic-interaction chromatography. The partial amino-acid sequences reflect high similarities between the four proteins as well as with the sequences established for carrot, spinach, millet and maize nonspecific lipid-transfer proteins. All these sequences show conservation of three cysteines at positions 4, 14 and 30-32, as well as glycine, valine, tyrosine and lysine residues at positions 5, 7, 17 and 37, respectively. In-vitro lipid-transfer assays reveal that the four proteins catalyze the transfer of phosphatidylcholine from liposomes towards mitochondria with an efficiency similar or higher than that of a purified maize lipid-transfer protein.

Amino Acid Sequence↗
PubMed1993

Acyl-binding/lipid-transfer proteins from rape seedlings, a novel category of proteins interacting with lipids.

From rape (Brassica napus) seedlings proteins able to bind fatty acids and their CoA-esters were purified by gel filtration and cation-exchange chromatography. Among the four proteins detected, one of them (peak IV) appeared purified to homogeneity. This protein is a monomer with a molecular mass of about 9 kDa, as estimated by gel filtration and by polyacrylamide gel electrophoresis. The isoelectric point of the rape protein was higher than 10.5 as determined by chromatofocusing. The pure rape protein appeared furthermore to be able to transfer several phospholipids (phosphatidylcholine, phosphatidylinositol and phosphatidylethanolamine) between membranes. The rape protein, having a multifunctional property, was thus called acyl-binding/lipid-transfer protein (AB-LTP). In order to compare this protein to plant lipid-transfer proteins (LTPs), its structure was determined. The amino acid analysis of the rape AB-LTP revealed a high amount of alanine, an absence of histidine and tryptophan and the presence of eight cysteine residues. The N-terminal amino acid sequence of the rape protein revealed a high homology to plant LTPs. These observations led us to propose that the rape AB-LTPs belong to a category of plant proteins interacting with lipids and playing a role in the fatty acid dynamics.

Amino Acid Sequence↗
PubMed1993

Control of maize lipid transfer protein activity by oxido-reducing conditions.

The activity of lipid transfer proteins (LTPs) isolated from maize, able to facilitate phospholipid movement between membranes, was studied under various oxido-reducing conditions. A progressive inactivation of LTP transfer activity was observed with increasing concentrations of reduced dithiothreitol (DTTred). This inactivation was accompanied by an increase in SH titer as well as by changes of the protein conformation deduced from its higher mobility in SDS-PAGE. By contrast, DTTred did not affect the formation of lipid-LTP complex. Transfer activity and original electrophoretic mobility were partially restored under reoxidation by air or oxidized DTT. Together, these results demonstrate the critical role of correct S-S bondings on LTP activity and suggest a possible in vivo regulation, according to the specific oxido-reducing conditions prevailing in different cell compartments.

Antigens, Plant↗
PubMed1993

Biosynthesis of gamma-linolenic acid in developing seeds of borage (Borago officinalis L.).

delta 6-desaturation of [14C]linoleoyl-CoA or [14C]oleoyl-CoA leading to the synthesis of gamma-linolenic acid was studied in vitro with microsomal fractions from developing seeds of Borago officinalis. Time course of the reaction, effects of protein and precursor concentrations and nucleotide requirements were examined. These parameters allowed us to improve the in vitro delta 6-desaturation assay. We observed that the precursors were acylated mainly in phosphatidylcholine, diacylglycerol and triacylglycerol, and then desaturated. NADH was absolutely required when [14C]oleoyl-CoA was the precursor, but not when [14C]linoleoyl-CoA was the precursor although it stimulated the reaction. The in vitro delta 6-desaturase activity was found mainly in phosphatidylcholine, associated with enriched endoplasmic reticulum membranes (ER) from embryos. No activity was observed in ER from seed coat or seedling. During maturation of the seeds, delta 6-desaturase reached its highest activity 14 to 16 days after pollination.

Fatty Acid Desaturases↗
PubMed1992

Transmembrane redistribution of phospholipids of the human red cell membrane during hypotonic hemolysis.

The transmembrane distribution of spin-labeled phospholipids was measured in human erythrocytes before and after hypotonic hemolysis by electron paramagnetic resonance. With a first series of partially water soluble probes a complete randomization of phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine and sphingomyelin analogues was achieved when cells were resealed in the absence of Mg-ATP or when the aminophospholipid translocase was inhibited by vanadate or calcium. If the ghosts were resealed with Mg-ATP inside, the transmembrane asymmetry of the aminophospholipids was reestablished. With long chain insoluble spin-labeled lipids complete randomization was obtained with the phosphatidylcholine analogue but even in the presence of vanadate only a small percentage (approx. 15%) of the spin-labeled phosphatidylserine flopped to the outer monolayer and comparable percentage of the spin-labeled sphingomyelin flipped to the inner monolayer, indicating a hierarchy in the phospholipid redistribution for these water insoluble lipids during hemolysis. The mechanism by which a selective randomization takes place is not known. It may involve phosphatidylserine-protein interactions in the inner leaflet and sphingomyelin-cholesterol or sphingomyelin-sphingomyelin interaction in the outer leaflet.

Electron Spin Resonance Spectroscopy↗
PubMed1991

Purification and some properties of the squalene-tetrahymanol cyclase from Tetrahymena thermophila.

The membrane-bound enzyme from Tetrahymena thermophila responsible for the conversion of squalene into the quasi-hopanoid tetrahymanol was purified 297-fold to near homogeneity. Purification involved solubilization by octylthioglucoside, chromatography on DEAE-trisacryl, hydroxyapatite and FPLC ion-exchange on Mono Q. The apparent KM was found to be 18 microM. 2,3-Iminosqualene and N,N-dimethyldodecylamine-N-oxide are effective inhibitors of the cyclase with I50 values of 50 and 30 nM, respectively. The cyclase has a molecular mass of 72 kDa as judged by electrophoresis in polyacrylamide gels under denaturating conditions. The optimal enzymatic activity was obtained at pH 7.0 and 30 degrees C. The solubilized enzyme needs the presence of detergent for maintaining activity. The influence of different detergents on cyclase activity was studied. Triton X-100 proved to be a strong inactivator of the enzyme. Solubilization of the cyclase in Tween 80 and digitonin inactivates the enzyme. However, its activity can be recovered by complementation of the assay buffer with octylthioglucoside above its critical micellar concentration. We suggest that this approach might be applicable to other membrane-bound proteins.

Animals↗
PubMed1991

Use of spin-labeled and fluorescent lipids to study the activity of the phospholipid transfer protein from maize seedlings.

The transfer of spin-labeled and fluorescent lipids between sonicated vesicles and different host membranes has been measured in the presence or absence of a phospholipid transfer protein purified from maize seedlings. It was found that the protein has little specificity towards the phospholipid head group and allows the transfer of hydrophobic long chain phospholipids. By contrast, no transfer of a cholesterol analogue could be detected. By EPR spectroscopy, evidence is presented that shows that the protein catalyzes the incorporation of labeled phospholipids in the outer monolayer of the acceptor membranes. The efficiency of the transfer depends largely on the nature of the acceptor: erythrocytes are more difficult to label than chromaffin granules or liposomes made with unsaturated lipids. Thus, consistent with the high activation energy observed, the transfer is facilitated when it involves fluid membranes. These results are in favor of a process involving the exchange of phospholipids, facilitated by a shuttle protein rather than a fusion mechanism.

Ascorbic Acid↗
PubMed1991

Multiple mRNA coding for phospholipid-transfer protein from Zea mays arise from alternative splicing.

We have isolated a novel cDNA coding for maize phospholipid-transfer protein. The cDNA sequence is similar to the first one obtained by Tchang et al. [J. Biol. Chem. 263 (1988) 16849-16855] differing only by a mslal number of nucleotide substitutions and insertions. One of these insertions is 74 bp long and is flanked by consensus intron splicing sequences. The protein coded by the two cDNA has identical amino acids except in the C terminus. This difference derived from the presence of the 74-bp insert. The possible existence of an alternative splicing mechanism that could introduce heterogeneity in the sequence of these proteins is proposed.

Amino Acid Sequence↗
PubMed1991

NADH-Ferricyanide Reductase of Leaf Plasma Membranes : Partial Purification and Immunological Relation to Potato Tuber Microsomal NADH-Ferricyanide Reductase and Spinach Leaf NADH-Nitrate Reductase.

Plasma membranes obtained by two-phase partitioning of microsomal fractions from spinach (Spinacea oleracea L. cv Medania) and sugar beet leaves (Beta vulgaris L.) contained relatively high NADH-ferricyanide reductase and NADH-nitrate reductase (NR; EC 1.6.6.1) activities. Both of these activities were latent. To investigate whether these activities were due to the same enzyme, plasma membrane polypeptides were separated with SDS-PAGE and analyzed with immunoblotting methods. Antibodies raised against microsomal NADH-ferricyanide reductase (tentatively identified as NADH-cytochrome b(5) reductase, EC 1.6.2.2), purified from potato (Solanum tuberosum L. cv Bintje) tuber microsomes, displayed one single band at 43 kilodaltons when reacted with spinach plasma membranes, whereas lgG produced against NR from spinach leaves gave a major band at 110 kilodaltons together with a few fainter bands of lower molecular mass. Immunoblotting analysis using inside-out and right-side-out plasma membrane vesicles strongly indicated that NR was not an integral protein but probably trapped inside the plasma membrane vesicles during homogenization. Proteins from spinach plasma membranes were solubilized with the zwitterionic detergent 3-[(3-cholamidopropyl) dimethylammonio] 1-propane-sulfonate and separated on a Mono Q anion exchange column at pH 5.6 with fast protein liquid chromatography. One major peak of NADH-ferricyanide reductase activity was found after separation. The peak fraction was enriched about 70-fold in this activity compared to the plasma membrane. When the peak fractions were analyzed with SDS-PAGE the NADH-ferricyanide reductase activity strongly correlated with a 43 kilodalton polypeptide which reacted with the antibodies against potato microsomal NADH-ferricyanide reductase. Thus, our data indicate that most, if not all, of the truly membrane-bound NADH-ferricyanide reductase activity of leaf plasma membranes is due to an enzyme very similar to potato tuber microsomal NADH-ferricyanide reductase (NADH-cytochrome b(5) reductase).

Journal Article↗
PubMed1990

Lipid transfer in plants.

Plant cells contain cytosolic proteins, called lipid transfer proteins (LTP), which are able to facilitate in vitro intermembrane transfer of phospholipids. Proteins of this kind from three plants, purified to homogeneity, have several properties in common: molecular mass around 9 kDa, high isoelectric point, lack of specificity for phospholipids, and binding ability for fatty acids. The comparison of their amino acid sequences revealed striking homologies and conserved domains which are probably involved in their function as LTPs. These proteins could play a major role in membrane biogenesis by conveying phospholipids from their site of biosynthesis to membranes unable to form these lipids. Immunochemical methods were used to establish an in vivo correlation between membrane biogenesis and the level of LTP or the amount of LTP synthesized in vitro from mRNAs. The recent isolation of a full-length cDNA allows novel approaches to studying the participation of LTPs in the biogenesis of plant cell membranes.

Amino Acid Sequence↗
PubMed1990

Purification and characterization of a novel phospholipid transfer protein from filamentous fungi.

1. We have isolated from mycelia of Mucor mucedo, a filamentous fungus, a phospholipid transfer protein. 2. The purification steps were gel filtration, hydroxyapatite chromatography, blue affinity column and fast protein liquid chromatography on anion exchanger. 3. A purified protein was obtained with a molecular mass of 24 kDa and a pI of 5.05 and its N-terminal sequence was established. 4. This protein transfers phosphatidylinositol, as well as phosphatidylcholine and phosphatidylethanolamine.

Amino Acid Sequence↗
PubMed1989

Changes in Level and Activity of Phospholipid Transfer Protein during Maturation and Germination of Maize Seeds.

The variations of the amounts of phospholipid transfer proteins (PLTP), determined by ELISA and immunoblotting methods, were followed during the maturation and germination of maize (Zea mays L.) seeds. Changes of the amounts of PLTP occur during seed maturation. The levels of PLTP, low in the first 3 weeks after fecondation, strongly raised 3 to 5 weeks after, then reached and maintained a high value (10% of total soluble proteins) during the last steps of maturation. These variations, determined by ELISA, are in accordance with the observations made by immunoblotting. Changes in phospholipid transfer activity were also found when protein extracts prepared from seeds at different stages of maturation were assayed for transfer activity. The levels of PLTP were also determined during the germination of maize seeds and the early growth of the plantlets, both in the endosperm and the aerial parts. While no major change was observed in the endosperm, a high increase in PLTP level was found in the aerial part of the plantlet, both by ELISA and immunoblotting. An enhancement of the phospholipid transfer activity was parallely observed in the protein extracts of plantlets at various stages of germination. These results are consistent with an in vivo correlation between the synthesis of phospholipid transfer protein, observed during the maturation and germination of maize seeds, and the biogenesis of membranes which involves intracellular movements of phospholipids.

Journal Article↗
PubMed1988

Synthesis of phospholipid transfer proteins from maize seedlings.

The synthesis of phospholipid transfer proteins has been studied in vitro after isolation of poly(A)+RNAs from maize seedlings and by in vivo labelling of coleoptiles. After immunoprecipitation of translation products in wheat germ or in reticulocyte lysate systems, the analysis by electrophoresis revealed two bands of molecular mass 9 kDa and 12 kDa. The in vitro synthesized 12 kDa protein is a precursor of the 9 kDa purified protein from maize seedlings as suggested by competition experiments with the pure protein. After immunoprecipitation of in vivo labelled proteins, two bands were detected. One of them, having a molecular mass of 7 kDa, could be related to the in vitro synthesized 9 kDa protein, the other corresponding to the purified protein. Furthermore, biosynthesis of both precursors occurs on membrane-bound polysomes. Presumably a post translational process occurs, yielding to the mature forms.

Carrier Proteins↗
PubMed1988

Hydrophobic-cluster analysis of plant protein sequences. A domain homology between storage and lipid-transfer proteins.

Hydrophobic-cluster analysis was used to characterize a conserved domain located near the C-terminal amino acid sequence of wheat (Triticum aestivum) storage proteins. This domain was transformed into a linear template for a global search for similarities in over 5200 protein sequences. In addition to proteins that had already been found to exhibit homology to wheat storage proteins, a previously unreported homology was found with non-specific lipid-transfer proteins from castor bean (Ricinus communis) and from spinach (Spinacia oleracea) leaf. Hydrophobic-cluster analysis of various members of the present protein group clearly shows a typical domain structure where (i) variable and conserved domains are located along the sequence at precise positions, (ii) the conserved domains probably reflect a common ancestor, and (iii) the unique properties of a given protein (chain cut into subunits, repetitive domains, trypsin-inhibitor active site) are associated with the variable domains.

Amino Acid Sequence↗