Purification and characterization of ten new rice NaCl-soluble proteins: identification of four protein-synthesis inhibitors and two immunoglobulin-binding proteins |
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Authors: | Gabriel G Limas Matilde Salinas Ignacio Moneo Stefan Fischer Brigitte Wittmann-Liebold Enrique Méndez |
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Institution: | (1) Servicio de Endocrinologia, Hospital Ramón y Cajal , E-28034 Madrid, Spain;(2) Servicio de Bioquimica, Hospital Ramón y Cajal , E-28034 Madrid, Spain;(3) Servicio de Inmunologia, Hospital Ramón y Cajal , E-28034 Madrid, Spain;(4) Knauer Company, Hegauer Weg 38, D-1000 Berlin 37;(5) Max-Planck-Institut für Molekulare Genetik, Ihnestrasse 73, D-1000 Berlin 33 |
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Abstract: | Ten new proteins from rice (Oryza saliva L. cv. Bahia) including four protein-synthesis inhibitors and two immunoglobulin E (IgE)-binding proteins have been isolated and characterized. These proteins as well as one previously known component, -globulin, were purified from a 0.5 M NaCl extract of rice endosperm by a new, apparently non-denaturing, isolation procedure developed for rice proteins. The method is based on extractions of this complex protein mixture with a diluted volatile salt solution and an aqueous solution of ethanol. This preliminary step results in an improvement in the separation of these proteins, thus facilitating their subsequent purification by reversed-phased high-performance liquid chromatography. These new proteins have similar relative molecular masses (Mrs) from 11000 to 17000. The purity of the proteins was analyzed by micro two-dimensional gel electrophoresis. Four of these components were found to be in-vitro protein-synthesis inhibitors in a cell-free system from rat brain. The NH2-terminal amino-acid sequences of these four inhibitors were determined from 12 to 26 cycles after direct blotting of the separated proteins from electrophoresis gels. Three of these proteins with Mrs between 16000 and 17000 showed a high degree of homology ranging from 57% to 75% but seem to be unrelated to the fourth inhibitor. In addition, the -globulin and one of the new low-molecular-weight proteins of Mr 12500 seemed to show allergenic properties since they bound IgE antibodies from the sera of hypersensitive patients. Boths proteins have blocked NH2-terminal amino acids.Abbreviations HMW
high molecular weight
- IgE
immunoglobulin E
- LMW
low molecular weight
- Mr
relative molecular mass
- PAGE
polyacrylamide gel electrophoresis
- RP-HPLC
reversed-phase high-performance liquid chromatography
- SDS
sodium dodecyl sulphate
We thank F. Soriano and F. Colillia for technical assistance, and Shirley McGrath for secretarial work. We also appreciate the cheerful assistance of the members of Instituto Nacional de Semillas, specially Mr. L. Solaices, who provided samples of rice. This work was supported by a grant from Comision Asesora de Investigación Ciéntifica y Técnica. |
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Keywords: | Allergenic protein Oryza (proteins) Protein (NaCl-soluble) Protein synthesis inhibitor Sodium chloride-soluble protein |
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