Tarallo Mariana, Hayhurst Max, Loo Trevor S, Gerth Monica L, Bradshaw Rosie E
Bioprotection Aotearoa, School of Food Technology and Natural Sciences, Massey University, Palmerston North, New Zealand.
School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand.
Methods Mol Biol. 2025;2892:233-248. doi: 10.1007/978-1-0716-4330-3_16.
This chapter describes the protocol for heterologous expression of Phytophthora proteins in the yeast Pichia pastoris. Two methods to prepare the constructs for expression are described, using two different strains of P. pastoris, as well as methods for protein expression and purification by immobilized metal ion affinity (IMAC). The combined protocol was used to successfully produce a Phytophthora pluvialis small, secreted cell death elicitor protein and a Phytophthora agathidicida glycoside hydrolase 12, PaXEG1, also a cell death-inducing protein.
本章描述了疫霉属蛋白在巴斯德毕赤酵母中进行异源表达的实验方案。文中介绍了使用两种不同的巴斯德毕赤酵母菌株制备表达构建体的两种方法，以及通过固定化金属离子亲和色谱（IMAC）进行蛋白表达和纯化的方法。该组合方案已成功用于生产一种来自雨生疫霉的小分泌型细胞死亡诱导蛋白，以及一种来自阿加西疫霉的糖苷水解酶12（PaXEG1），它也是一种细胞死亡诱导蛋白。
