Assembly and purification of AvrSr35-induced Sr35 resistosome and determination of its structure by cryo-EM

Meng-Xi Liu; Yan-Bo Zhao; Ze-Kai Li; Tao-Tao Chen; Songying Ouyang

STAR Protocols (Dec 2022)

Assembly and purification of AvrSr35-induced Sr35 resistosome and determination of its structure by cryo-EM

Meng-Xi Liu,
Yan-Bo Zhao,
Ze-Kai Li,
Tao-Tao Chen,
Songying Ouyang

Affiliations

Meng-Xi Liu: Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, the Key Laboratory of Innate Immune Biology of Fujian Province, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China
Yan-Bo Zhao: Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, the Key Laboratory of Innate Immune Biology of Fujian Province, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China
Ze-Kai Li: Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, the Key Laboratory of Innate Immune Biology of Fujian Province, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China
Tao-Tao Chen: Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, the Key Laboratory of Innate Immune Biology of Fujian Province, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China
Songying Ouyang: Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, the Key Laboratory of Innate Immune Biology of Fujian Province, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China; Corresponding author

Journal volume & issue: Vol. 3, no. 4
p. 101796

Abstract

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Summary: Sr35, a coiled-coil nucleotide-binding leucine-rich repeat receptor (CC-NLR) from the wheat species Triticum monococcum can directly recognize the pathogen avirulence factor AvrSr35 and confers immunity against wheat stem rust caused by Puccinia graminis f.sp. tritici race Ug99. Assembly of a stable Sr35 resistosome induced by AvrSr35 in vitro is usually limited by protein expression and low assembly efficiency. Here, we describe the expression and purification of AvrSr35 and Sr35, in vitro assembly of Sr35 resistosome for structure determination by cryo-EM.For complete details on the use and execution of this protocol, please refer to Zhao et al. (2022). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Published in STAR Protocols

ISSN: 2666-1667 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science: Science (General)
Website: https://star-protocols.cell.com/

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