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Gel filtration is frequently used to study the behaviour and composition of protein complexes. In previous studies, gel filtration analysis of solubilised membranes containing the tomato Cf-4 and Cf-9 resistance proteins indicated that these Cf proteins are present in an approximately 400- and 420-kDa protein complex, respectively, which contains only one Cf molecule per complex, does not contain Rho-related proteins, and does not alter in size upon elicitation. Here, we show that inactive Cf-4 and Cf-9 mutant proteins have a similar large apparent size upon gel filtration analysis. The size remains unaltered after pre-treating the samples under harsh conditions, such as boiling with SDS and incubation in 6 m urea. A similar large apparent size was found for Cf-4 and Cf-9 isolated from SDS gel and for Cf-9 expressed by insect cells. Therefore, the large apparent size observed in our studies appears to be an intrinsic property of the Cf proteins, rather than being caused by association with high-molecular-weight protein(s). Taken together, these results suggest that caution should be taken when interpreting data obtained from gel filtration of LRR-containing proteins.

Original publication

DOI

10.1046/j.1365-313X.2003.01803.x

Type

Journal article

Journal

Plant J

Publication Date

08/2003

Volume

35

Pages

305 - 315

Keywords

Animals, Base Sequence, Cell Line, Chromatography, Gel, DNA, Plant, Detergents, Genetic Vectors, Glycosylation, Macromolecular Substances, Membrane Glycoproteins, Molecular Weight, Mutation, Plant Proteins, Plants, Genetically Modified, Recombinant Proteins, Rhizobium, Solubility, Tobacco