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Review

Chromatography-Free Purification Strategies for Large Biological Macromolecular Complexes Involving Fractionated PEG Precipitation and Density Gradients

1
Department of Structural Dynamics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany
2
Research Group for Structural Biochemistry and Mechanisms, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany
*
Author to whom correspondence should be addressed.
Academic Editors: Augusto Quaresma Henriques Pedro, Mara G. Freire, Fani Pereira de Sousa and Patrícia Alexandra Nunes Pereira
Life 2021, 11(12), 1289; https://doi.org/10.3390/life11121289
Received: 9 October 2021 / Revised: 10 November 2021 / Accepted: 22 November 2021 / Published: 24 November 2021
A complex interplay between several biological macromolecules maintains cellular homeostasis. Generally, the demanding chemical reactions which sustain life are not performed by individual macromolecules, but rather by several proteins that together form a macromolecular complex. Understanding the functional interactions amongst subunits of these macromolecular machines is fundamental to elucidate mechanisms by which they maintain homeostasis. As the faithful function of macromolecular complexes is essential for cell survival, their mis-function leads to the development of human diseases. Furthermore, detailed mechanistic interrogation of the function of macromolecular machines can be exploited to develop and optimize biotechnological processes. The purification of intact macromolecular complexes is an essential prerequisite for this; however, chromatographic purification schemes can induce the dissociation of subunits or the disintegration of the whole complex. Here, we discuss the development and application of chromatography-free purification strategies based on fractionated PEG precipitation and orthogonal density gradient centrifugation that overcomes existing limitations of established chromatographic purification protocols. The presented case studies illustrate the capabilities of these procedures for the purification of macromolecular complexes. View Full-Text
Keywords: protein purification; PEG precipitation; protein precipitation; chromatography-free protein purification; sucrose density gradient centrifugation; 20S proteasome; 26S proteasome; fatty acid synthase; immunoproteasome; buffer optimization; monolithic columns; macromolecular complexes protein purification; PEG precipitation; protein precipitation; chromatography-free protein purification; sucrose density gradient centrifugation; 20S proteasome; 26S proteasome; fatty acid synthase; immunoproteasome; buffer optimization; monolithic columns; macromolecular complexes
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MDPI and ACS Style

Henneberg, F.; Chari, A. Chromatography-Free Purification Strategies for Large Biological Macromolecular Complexes Involving Fractionated PEG Precipitation and Density Gradients. Life 2021, 11, 1289. https://doi.org/10.3390/life11121289

AMA Style

Henneberg F, Chari A. Chromatography-Free Purification Strategies for Large Biological Macromolecular Complexes Involving Fractionated PEG Precipitation and Density Gradients. Life. 2021; 11(12):1289. https://doi.org/10.3390/life11121289

Chicago/Turabian Style

Henneberg, Fabian, and Ashwin Chari. 2021. "Chromatography-Free Purification Strategies for Large Biological Macromolecular Complexes Involving Fractionated PEG Precipitation and Density Gradients" Life 11, no. 12: 1289. https://doi.org/10.3390/life11121289

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