ABSTRACT
ß-1,3-glucanases are found in organisms as diverse as plants, animals, bacteria and fungi. In plants, such enzymes are not only associated with defense mechanisms against pathogens, but also play critical roles in physiological and developmental processes. Here we identified a new ß-1,3-glucanase in maize seeds, and named it ZmGlucA. Sequence analysis revealed that ZmGlucA belongs to the class A of ß-1,3-glucanase, a class related to defense and physiological processes in plants. mRNA and protein assays showed that zmGlucA is expressed exclusively in seeds, and it is differentially regulated during seed development. Additionally, zmGlucA expression is strongly induced in seeds of the mutant dek 827Kpro1, which is defective for embryo and endosperm development. Our data support the idea that ZmGlucA protein is relevant to seed development.
Subject(s)
Glycoside Hydrolases/metabolism , Plants, Genetically Modified/growth & development , Seeds/enzymology , Seeds/growth & development , Zea mays/enzymology , Zea mays/growth & development , Blotting, Northern , Gene Expression Regulation, Enzymologic/genetics , Gene Expression Regulation, Enzymologic/physiology , Gene Expression Regulation, Plant/genetics , Gene Expression Regulation, Plant/physiology , Glycoside Hydrolases/genetics , Plants, Genetically Modified/genetics , Seeds/genetics , Zea mays/geneticsABSTRACT
The SALT protein is a 14.5 kDa mannose-binding lectin, originally described as preferentially expressed in rice plant roots in response to NaCl stress. Recombinant SALT lectin was produced in Escherichia coli from a cDNA clone encoding protein. After isopropyl-beta-d-thiogalactopyranoside induction, the expression level achieved was 23% of the soluble protein. The recombinant agglutinin was purified by a single-step process by dialyses against a high concentrated salt solution. After purification, hemagglutination assays of rabbit erythrocytes revealed that the recombinant SALT protein is a potent agglutinin (0.078 microg ml(-1) minimal concentration). The purified recombinant lectin was also used for comparative estimation of native protein amounts in protein extracts from rice plants by Western blot assay.
Subject(s)
Oryza/metabolism , Plant Proteins/biosynthesis , Plant Proteins/isolation & purification , Animals , Electrophoresis, Polyacrylamide Gel/methods , Erythrocytes/drug effects , Escherichia coli/genetics , Escherichia coli/metabolism , Hemagglutination Tests/methods , Oryza/genetics , Plant Lectins/biosynthesis , Plant Lectins/genetics , Plant Lectins/immunology , Plant Lectins/isolation & purification , Plant Proteins/genetics , Plant Proteins/immunology , Rabbits , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Recombinant Proteins/isolation & purificationABSTRACT
A practical and low cost system for isoelectric protein focusing (IEF) was developed. The system uses a multi-cell glass plate compatible with a common vertical one-dimensional electrophoresis chamber, dispensing specific IEF apparatus. After focusing, the IEF gels are easily recovered. The resulting two-dimensional (2-D) gels have provided efficient protein separation for different concentrations and extracts.
