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Crystallization of the rice immune receptor RGA5A_S with the rice blast fungus effector AVR1‐CO39 prepared via mixture and tandem strategies
Author(s) -
Guo Liwei,
Zhang Yikun,
Ma Mengqi,
Liu Qiang,
Zhang Yanan,
Peng Youliang,
Liu Junfeng
Publication year - 2018
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x18003618
Subject(s) - crystallization , crystallography , effector , molecule , monoclinic crystal system , oryza sativa , tartrate , chemistry , crystal (programming language) , stereochemistry , crystal structure , biochemistry , organic chemistry , computer science , programming language , gene
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 P 3 1 21, 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 I 4, 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 two molecules of AVR1‐CO39 in the asymmetric unit of the initial model. Further structural characterization of the interaction between RGA5A_S and AVR1‐CO39 will lead to a better understanding of the mechanism underlying effector recognition by R proteins.