Crystallization of the rice immune receptor RGA5A_S with the rice blast fungus effector AVR1-CO39 prepared via mixture and tandem strategies

RGA5 is a component of the Pia resistance-protein pair (RGA4/RGA5) from Oryza sativa L. japonica. It acts as an immune receptor that directly recognizes the effector AVR1-CO39 from Magnaporthe oryzae via a C-terminal non-LRR domain (RGA5A_S). The interaction between RGA5A_S and AVR1-CO39 relieves the repression of RGA4, leading to effector-independent cell death. To determine the structure of the complex of RGA5A_S and AVR1-CO39 and to understand the details of this interaction, the complex was prepared by fusing the proteins together, by mixing them in vitro or by co-expressing them in one host cell. Samples purified via the first two strategies were crystallized under two different conditions. A mixture of AVR1-CO39 and RGA5A_S (complex I) crystallized in 1.1   M ammonium tartrate dibasic, 0.1   M sodium acetate – HCl pH 4.6, while crystals of the fusion complex RGA5A_S-TEV-AVR1-CO39 (complex II) were grown in 2   M NaCl. The crystal of complex I belonged to space group P3121, with unit-cell parameters a   =   b = 66.2, c = 108.8   Å , α = β = 90, γ = 120 ° . The crystals diffracted to a Bragg spacing of 2.4   Å , and one molecule each of RGA5A_S and AVR1-CO39 were present in the asymmetric unit of the initial model. The crystal of complex II belonged to space group I4, with unit-cell parameters a = b = 137.4, c = 66.2   Å , α = β = γ = 90 ° . The crystals diffracted to a Bragg spacing of 2.72   Å , and there were two molecules of RGA5A_S and t...

Source: Acta Crystallographica Section F - March 28, 2018 Category: Biochemistry Authors: Guo, L. Zhang, Y. Ma, M. Liu, Q. Zhang, Y. Peng, Y. Liu, J. Tags: RGA5A_S AVR1-CO39 rice Magnaporthe oryzae resistance protein Avr effector crystal complex research communications Source Type: research