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Copurification of rho protein and the rho-GDP dissociation inhibitor from bovine neutrophil cytosol. Effect of phosphoinositides on rho ADP-ribosylation by the C3 exoenzyme of Clostridium botulinum. 1992

N Bourmeyster, and M J Stasia, and J Garin, and J Gagnon, and P Boquet, and P V Vignais
Laboratoire de Biochimie, Centre d'Etudes Nucléaires, Grenoble, France.

The substrate of the C3 exoenzyme from botulinum toxin is a protein which is particularly abundant in the cytosol of neutrophils [Stasia, M. J., Jouan, A., Bourmeyster, N., Boquet, P., & Vignais, P. V. (1991) Biochem. Biophys. Res. Commun. 180, 615-622]. Optimal conditions for the ADP-ribosylation of the C3 substrate have been established in order to follow the course of its purification from bovine neutrophil cytosol. In particular, phosphoinositides at micromolar concentrations were found to enhance the ADP-ribosylation capacity of the C3 substrate in crude neutrophil cytosol and partially purified fractions. A [32P]ADP-ribosylatable protein, migrating on SDS-PAGE with a mass of 24 kDa, was copurified with a 29-kDa protein by a series of chromatographic steps on DEAE-Sephacel, Biogel P60, and Mono Q. In the case of the C3 substrate, isoelectric focusing revealed two major labeled bands with pI values of 6.2 and 5.6; the pI of the 29-kDa protein was 4.8-5.0. On the basis of the amino acid sequence of peptides resolved after proteolytic digestion, the 24-kDa protein and the 29-kDa protein were identified respectively as rho and the GDP dissociation inhibitor (GDI), suggesting that rho and GDI copurify from bovine neutrophil cytosol in the form of a complex. The presence of a number of amino acid residues specific of rho A in the enzymatic digest originating from rho indicates that, among the rho proteins, at least rho A belongs to the GDI-rho complex.

UI MeSH Term Description Entries
D008565 Membrane Proteins Proteins which are found in membranes including cellular and intracellular membranes. They consist of two types, peripheral and integral proteins. They include most membrane-associated enzymes, antigenic proteins, transport proteins, and drug, hormone, and lectin receptors. Cell Membrane Protein,Cell Membrane Proteins,Cell Surface Protein,Cell Surface Proteins,Integral Membrane Proteins,Membrane-Associated Protein,Surface Protein,Surface Proteins,Integral Membrane Protein,Membrane Protein,Membrane-Associated Proteins,Membrane Associated Protein,Membrane Associated Proteins,Membrane Protein, Cell,Membrane Protein, Integral,Membrane Proteins, Integral,Protein, Cell Membrane,Protein, Cell Surface,Protein, Integral Membrane,Protein, Membrane,Protein, Membrane-Associated,Protein, Surface,Proteins, Cell Membrane,Proteins, Cell Surface,Proteins, Integral Membrane,Proteins, Membrane,Proteins, Membrane-Associated,Proteins, Surface,Surface Protein, Cell
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D008970 Molecular Weight The sum of the weight of all the atoms in a molecule. Molecular Weights,Weight, Molecular,Weights, Molecular
D009504 Neutrophils Granular leukocytes having a nucleus with three to five lobes connected by slender threads of chromatin, and cytoplasm containing fine inconspicuous granules and stainable by neutral dyes. LE Cells,Leukocytes, Polymorphonuclear,Polymorphonuclear Leukocytes,Polymorphonuclear Neutrophils,Neutrophil Band Cells,Band Cell, Neutrophil,Cell, LE,LE Cell,Leukocyte, Polymorphonuclear,Neutrophil,Neutrophil Band Cell,Neutrophil, Polymorphonuclear,Polymorphonuclear Leukocyte,Polymorphonuclear Neutrophil
D010716 Phosphatidylinositols Derivatives of phosphatidic acids in which the phosphoric acid is bound in ester linkage to the hexahydroxy alcohol, myo-inositol. Complete hydrolysis yields 1 mole of glycerol, phosphoric acid, myo-inositol, and 2 moles of fatty acids. Inositide Phospholipid,Inositol Phosphoglyceride,Inositol Phosphoglycerides,Inositol Phospholipid,Phosphoinositide,Phosphoinositides,PtdIns,Inositide Phospholipids,Inositol Phospholipids,Phosphatidyl Inositol,Phosphatidylinositol,Inositol, Phosphatidyl,Phosphoglyceride, Inositol,Phosphoglycerides, Inositol,Phospholipid, Inositide,Phospholipid, Inositol,Phospholipids, Inositide,Phospholipids, Inositol
D001905 Botulinum Toxins Toxic proteins produced from the species CLOSTRIDIUM BOTULINUM. The toxins are synthesized as a single peptide chain which is processed into a mature protein consisting of a heavy chain and light chain joined via a disulfide bond. The botulinum toxin light chain is a zinc-dependent protease which is released from the heavy chain upon ENDOCYTOSIS into PRESYNAPTIC NERVE ENDINGS. Once inside the cell the botulinum toxin light chain cleaves specific SNARE proteins which are essential for secretion of ACETYLCHOLINE by SYNAPTIC VESICLES. This inhibition of acetylcholine release results in muscular PARALYSIS. Botulin,Botulinum Neurotoxin,Botulinum Neurotoxins,Clostridium botulinum Toxins,Botulinum Toxin,Neurotoxin, Botulinum,Neurotoxins, Botulinum,Toxin, Botulinum,Toxins, Botulinum,Toxins, Clostridium botulinum
D002417 Cattle Domesticated bovine animals of the genus Bos, usually kept on a farm or ranch and used for the production of meat or dairy products or for heavy labor. Beef Cow,Bos grunniens,Bos indicus,Bos indicus Cattle,Bos taurus,Cow,Cow, Domestic,Dairy Cow,Holstein Cow,Indicine Cattle,Taurine Cattle,Taurus Cattle,Yak,Zebu,Beef Cows,Bos indicus Cattles,Cattle, Bos indicus,Cattle, Indicine,Cattle, Taurine,Cattle, Taurus,Cattles, Bos indicus,Cattles, Indicine,Cattles, Taurine,Cattles, Taurus,Cow, Beef,Cow, Dairy,Cow, Holstein,Cows,Dairy Cows,Domestic Cow,Domestic Cows,Indicine Cattles,Taurine Cattles,Taurus Cattles,Yaks,Zebus
D002851 Chromatography, High Pressure Liquid Liquid chromatographic techniques which feature high inlet pressures, high sensitivity, and high speed. Chromatography, High Performance Liquid,Chromatography, High Speed Liquid,Chromatography, Liquid, High Pressure,HPLC,High Performance Liquid Chromatography,High-Performance Liquid Chromatography,UPLC,Ultra Performance Liquid Chromatography,Chromatography, High-Performance Liquid,High-Performance Liquid Chromatographies,Liquid Chromatography, High-Performance
D003600 Cytosol Intracellular fluid from the cytoplasm after removal of ORGANELLES and other insoluble cytoplasmic components. Cytosols
D004591 Electrophoresis, Polyacrylamide Gel Electrophoresis in which a polyacrylamide gel is used as the diffusion medium. Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs

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