Open AccessThe Tomato R Gene Products I-2 and Mi-1 Are Functional ATP Binding Proteins with ATPase Activity
Author(s) -
W.I.L. Tameling,
S.D.J. Elzinga,
Patricia S. Darmin,
Jack H. Vossen,
Frank L. W. Takken,
Michel A. Haring,
Ben J. C. Cornelissen
Publication year - 2002
Publication title -
the plant cell
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.324
H-Index - 341
eISSN - 1532-298X
pISSN - 1040-4651
DOI - 10.1105/tpc.005793
Subject(s) - biology , biochemistry , nucleotide , binding site , mutant , gtp' , binding domain , gene , microbiology and biotechnology , atpase , nucleoside diphosphate kinase , kinase , enzyme
Most plant disease resistance (R) genes known today encode proteins with a central nucleotide binding site (NBS) and a C-terminal Leu-rich repeat (LRR) domain. The NBS contains three ATP/GTP binding motifs known as the kinase-1a or P-loop, kinase-2, and kinase-3a motifs. In this article, we show that the NBS of R proteins forms a functional nucleotide binding pocket. The N-terminal halves of two tomato R proteins, I-2 conferring resistance to Fusarium oxysporum and Mi-1 conferring resistance to root-knot nematodes and potato aphids, were produced as glutathione S-transferase fusions in Escherichia coli. In a filter binding assay, purified I-2 was found to bind ATP rather than other nucleoside triphosphates. ATP binding appeared to be fully dependent on the presence of a divalent cation. A mutant I-2 protein containing a mutation in the P-loop showed a strongly reduced ATP binding capacity. Thin layer chromatography revealed that both I-2 and Mi-1 exerted ATPase activity. Based on the strong conservation of NBS domains in R proteins of the NBS-LRR class, we propose that they all are capable of binding and hydrolyzing ATP.