19 F Dark-State Exchange Saturation Transfer NMR Reveals Reversible Formation of Protein-Specific Large Clusters in High-Concentration Protein Mixtures

Analytical Chemistry
John M EdwardsAlexander P Golovanov

Abstract

Proteins frequently exist as high-concentration mixtures, both in biological environments and increasingly in biopharmaceutical co-formulations. Such crowded conditions promote protein-protein interactions, potentially leading to formation of protein clusters, aggregation, and phase separation. Characterizing these interactions and processes in situ in high-concentration mixtures is challenging due to the complexity and heterogeneity of such systems. Here we demonstrate the application of the dark-state exchange saturation transfer (DEST) NMR technique to a mixture of two differentially 19F-labeled 145 kDa monoclonal antibodies (mAbs) to assess reversible temperature-dependent formation of small and large protein-specific clusters at concentrations up to 400 mg/mL. 19F DEST allowed quantitative protein-specific characterization of the cluster populations and sizes for both mAbs in the mixture under a range of conditions. Additives such as arginine glutamate and NaCl also had protein-specific effects on the dark-state populations and cluster characteristics. Notably, both mAbs appear to largely exist as separate self-associated clusters, which mechanistically respond differently to changes in solution conditions. We show that fo...Continue Reading

References

Jun 27, 1988·Physical Review Letters·M Medina-Noyola
Nov 4, 2000·Journal of Biomolecular NMR·M HelgstrandP Allard
Jul 22, 2004·Journal of the American Chemical Society·Alexander P GolovanovLu-Yun Lian
Nov 27, 2004·Nature·Anna StradnerPeter Schurtenberger
Jul 8, 2010·Journal of the American Chemical Society·Nicolas L FawziG Marius Clore
Dec 7, 2011·Chemical Communications : Chem Comm·Juan A AguilarGareth A Morris
Dec 15, 2011·Methods in Molecular Biology·Arpana DuttaJudith Klein-Seetharaman
Jan 10, 2012·International Journal of Pharmaceutics·John G ElvinChristopher F van der Walle
Jul 24, 2012·Archives of Biochemistry and Biophysics·Søren K RasmussenTorben P Frandsen
May 8, 2013·Expert Opinion on Biological Therapy·Tadeusz Robak
Jun 6, 2014·Chemical Reviews·Francois-Xavier TheilletPhilipp Selenko
Jul 6, 2014·International Journal of Pharmaceutics·Priscilla KheddoAlexander P Golovanov
Oct 8, 2014·Annual Review of Cell and Developmental Biology·Anthony A HymanFrank Jülicher
Dec 17, 2014·International Journal of Molecular Sciences·Irina M KuznetsovaVladimir N Uversky
Dec 23, 2014·MAbs·Dawn M EckerHoward L Levine
Feb 25, 2015·Quarterly Reviews of Biophysics·Nicholas J Anthis, G Marius Clore
Apr 18, 2015·The Journal of Physical Chemistry. B·Gregory V BarnettChristopher J Roberts
Aug 10, 2015·Analytical and Bioanalytical Chemistry·Anupama RanganathanBruce D Hammock
May 18, 2016·Biomaterials·Karl Fischer, Manfred Schmidt
Jul 12, 2017·European Journal of Pharmaceutics and Biopharmaceutics : Official Journal of Arbeitsgemeinschaft Für Pharmazeutische Verfahrenstechnik E.V·Patrick GaridelMichaela Blech
Jan 18, 2018·Journal of Pharmaceutical Sciences·Michael R Turner, Sathy V Balu-Iyer
Jun 5, 2018·Molecular Pharmaceutics·John M EdwardsAlexander P Golovanov

❮ Previous
Next ❯

Citations

Dec 31, 2020·International Journal of Molecular Sciences·Yuji Tokunaga, Koh Takeuchi
Apr 14, 2020·European Journal of Pharmaceutics and Biopharmaceutics : Official Journal of Arbeitsgemeinschaft Für Pharmazeutische Verfahrenstechnik E.V·Matja ZalarAlexander P Golovanov
Mar 5, 2021·ACS Pharmacology & Translational Science·Jack E BramhamAlexander P Golovanov

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.