European Molecular Biology Laboratory (EMBL), Meyerhofstrasse 1, 69117, Heidelberg, Germany.
Protein Purification Facility, The Wolfson Centre for Applied Structural Biology, The Hebrew University of Jerusalem, 91904, Jerusalem, Israel.
Microb Cell Fact. 2022 Apr 7;21(1):52. doi: 10.1186/s12934-022-01778-5.
Proteins are used as reagents in a broad range of scientific fields. The reliability and reproducibility of experimental data will largely depend on the quality of the (recombinant) proteins and, consequently, these should undergo thorough structural and functional controls. Depending on the downstream application and the biochemical characteristics of the protein, different sets of specific features will need to be checked.
A number of examples, representative of recurrent issues and previously published strategies, has been reported that illustrate real cases of recombinant protein production in which careful strategy design at the start of the project combined with quality controls throughout the production process was imperative to obtain high-quality samples compatible with the planned downstream applications. Some proteins possess intrinsic properties (e.g., prone to aggregation, rich in cysteines, or a high affinity for nucleic acids) that require certain precautions during the expression and purification process. For other proteins, the downstream application might demand specific conditions, such as for proteins intended for animal use that need to be endotoxin-free.
This review has been designed to act as a practical reference list for researchers who wish to produce and evaluate recombinant proteins with certain specific requirements or that need particular care for their preparation and storage.
蛋白质被广泛应用于各科学领域的试剂。实验数据的可靠性和可重复性在很大程度上取决于（重组）蛋白质的质量，因此应对这些蛋白质进行彻底的结构和功能控制。根据下游应用和蛋白质的生化特性，需要检查不同的特定特征集。
报告了一些代表性的实例，这些实例说明了重组蛋白生产中反复出现的问题和以前发表的策略，在项目开始时精心设计策略，并在整个生产过程中进行质量控制，对于获得与计划的下游应用兼容的高质量样品至关重要。一些蛋白质具有内在特性（例如易于聚集、富含半胱氨酸或与核酸有高亲和力），这在表达和纯化过程中需要特别注意。对于其他蛋白质，下游应用可能需要特定的条件，例如用于动物的蛋白质需要无内毒素。
本文旨在为希望生产和评估具有特定要求或需要特别注意其制备和储存的重组蛋白质的研究人员提供实用的参考清单。
