Heterogeneity of Protein Environments in Frozen Solutions and in the Dried State

Сальникова, М.М.; Varshney, Dushyant; Shalaev, Evgenyi

doi:10.1007/978-1-4939-2383-0_2

Cited by 11 publications

(6 citation statements)

References 51 publications

(59 reference statements)

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“…As a material is frozen, growth of ice crystals results in increase in the concentration of all solutes present in the sample, including dissolved air gasses (predominantly nitrogen and oxygen). As these gasses become more concentrated, air bubbles form, creating additional pathways for surface-related protein instability due to newly formed solution/air interface ( 44 ). The appearance of air bubbles during freezing has been directly observed by optical microscopy ( 45 ) and indirectly by small-angle neutron scattering (SANS) ( 46 ).…”

Section: Interfacial Stress In Drug Substance Developmentmentioning

confidence: 99%

Interfacial Stress in the Development of Biologics: Fundamental Understanding, Current Practice, and Future Perspective

Li¹,

Krause

Chen

et al. 2019

AAPS J

Self Cite

110

107

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Biologic products encounter various types of interfacial stress during development, manufacturing, and clinical administration. When proteins come in contact with vapor–liquid, solid–liquid, and liquid–liquid surfaces, these interfaces can significantly impact the protein drug product quality attributes, including formation of visible particles, subvisible particles, or soluble aggregates, or changes in target protein concentration due to adsorption of the molecule to various interfaces. Protein aggregation at interfaces is often accompanied by changes in conformation, as proteins modify their higher order structure in response to interfacial stresses such as hydrophobicity, charge, and mechanical stress. Formation of aggregates may elicit immunogenicity concerns; therefore, it is important to minimize opportunities for aggregation by performing a systematic evaluation of interfacial stress throughout the product development cycle and to develop appropriate mitigation strategies. The purpose of this white paper is to provide an understanding of protein interfacial stability, explore methods to understand interfacial behavior of proteins, then describe current industry approaches to address interfacial stability concerns. Specifically, we will discuss interfacial stresses to which proteins are exposed from drug substance manufacture through clinical administration, as well as the analytical techniques used to evaluate the resulting impact on the stability of the protein. A high-level mechanistic understanding of the relationship between interfacial stress and aggregation will be introduced, as well as some novel techniques for measuring and better understanding the interfacial behavior of proteins. Finally, some best practices in the evaluation and minimization of interfacial stress will be recommended.

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Section: Interfacial Stress In Drug Substance Developmentmentioning

confidence: 99%

Interfacial Stress in the Development of Biologics: Fundamental Understanding, Current Practice, and Future Perspective

Li¹,

Krause

Chen

et al. 2019

AAPS J

Self Cite

110

107

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show abstract

“…The FCS can be located in the lamellae and veins between the ice crystals, (micro)pockets within the ice structure, or puddles and grain boundary grooves on the ice surface. When the mixture is cooled below the eutectic temperature (T eu ), the FCS may crystallize or vitrify (Salnikova et al, 2015;Bogdan et al, 2014;Imrichova et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

The morphology of ice and liquid brine in an environmental scanning electron microscope: a study of the freezing methods

Vetráková

Neděla²,

Runštuk³

et al. 2019

The Cryosphere

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Abstract. The microstructure of polycrystalline ice with a threading solution of brine controls its numerous characteristics, including the ice mechanical properties, ice–atmosphere interactions, sea ice albedo, and (photo)chemical behavior in and on the ice. Ice samples were previously prepared in laboratories in order to study various facets of ice–impurity interactions and (photo)reactions to model natural ice–impurity behavior. We examine the impact of the freezing conditions and solute (CsCl used as a proxy for naturally occurring salts) concentrations on the microscopic structure of ice samples via an environmental scanning electron microscope. The method allows us to observe the ice surfaces in detail, namely, the free ice, brine puddles, brine-containing grain boundary grooves, individual ice crystals, and imprints left by entrapped air bubbles at temperatures higher than −25 ∘C. The amount of brine on the external surface is found proportional to the solute concentration and is strongly dependent on the sample preparation method. Time-lapse images in the condition of slight sublimation reveal subsurface association of air bubbles with brine. With rising temperatures (up to −14 ∘C), the brine surface coverage increases to remain enhanced during the subsequent cooling and until the final crystallization below the eutectic temperature. The ice recrystallization dynamics identify the role of surface spikes in retarding the ice boundaries' propagation (Zener pinning). The findings thus quantify the amounts of brine exposed to incoming radiation, available for the gas exchange, and influencing other mechanical and optical properties of ice. The results have straightforward and indirect implications for artificially prepared and naturally occurring salty ice, respectively.

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“…Further applications potentially include drug design, 17–18 dry powder inhaling, 19 studies of GPCR signaling mechanisms, 20 solid state NMR 21–23 and spin-label EPR 24–25 spectroscopy, and neutron scattering studies. 26 Here, we show the use of the powdered GPCR preparation method to investigate rhodopsin versus the ligand-free opsin apoprotein.…”

mentioning

confidence: 99%

Powdered G-Protein-Coupled Receptors

Perera

Chawla

Brown

2016

J. Phys. Chem. Lett.

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Preparation and storage of functional membrane proteins such as G-protein-coupled receptors (GPCRs) are crucial to the processes of drug delivery and discovery. Here we describe a method of preparing powdered GPCRs using rhodopsin as the prototype. We purified rhodopsin in CHAPS detergent with low detergent to protein ratio so the bulk of the sample represented protein (ca. 72% w/w). Our new method for generating powders of membrane proteins followed by rehydration paves the way for conducting functional and biophysical experiments. As an illustrative application powdered rhodopsin was prepared with and without the cofactor 11-cis retinal to enable partial rehydration of the protein with D2O in a controlled manner. Quasielastic neutron scattering studies using both spatial motion and energy landscape models form the basis for crucial insights into structural fluctuations and thermodynamics of GPCR activation.

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Heterogeneity of Protein Environments in Frozen Solutions and in the Dried State

Cited by 11 publications

References 51 publications

Interfacial Stress in the Development of Biologics: Fundamental Understanding, Current Practice, and Future Perspective

Interfacial Stress in the Development of Biologics: Fundamental Understanding, Current Practice, and Future Perspective

The morphology of ice and liquid brine in an environmental scanning electron microscope: a study of the freezing methods

Powdered G-Protein-Coupled Receptors

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