Salting out of Proteins Using Ammonium Sulfate Precipitation

Duong‐Ly, Krisna C.; Gabelli, Sandra B.

doi:10.1016/b978-0-12-420119-4.00007-0

Cited by 230 publications

(163 citation statements)

References 5 publications

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“…This ability of salt to completely shift the molecular driving forces of protein LLPS is consistent with the wide body of work demonstrating the significance of salt in the modulation of protein stability (40)(41)(42)(43), protein solubility (44,45), protein-protein interactions (46,47), and proteinnucleic acid interactions (48)(49)(50).…”

Section: Introductionsupporting

confidence: 86%

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

Krainer

Welsh

Joseph

et al. 2020

Preprint

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Many cellular proteins have the ability to demix spontaneously from solution to form liquid condensates. These phase-separated structures form membraneless compartments in living cells and have wide-ranging roles in health and disease. Elucidating the molecular driving forces underlying liquid-liquid phase separation (LLPS) of proteins has thus become a key objective for understanding biological function and malfunction. Here we show that proteins implicated in cellular phase separation, such as FUS, TDP-43, and Annexin A11, which form condensates at low salt concentrations via homotypic multivalent interactions, also have the ability to undergo LLPS at high salt concentrations by reentering into a phase-separated regime. Through a combination of experiments and simulations, we demonstrate that phase separation in the high-salt regime is mainly driven by hydrophobic and non-ionic interactions. As such, it is mechanistically distinct from the low-salt regime, where condensates are stabilized by a broad mix of electrostatic, hydrophobic, and non-ionic forces. Our work thus expands the molecular grammar of interactions governing LLPS of cellular proteins and provides a new view on hydrophobicity and non-ionic interactions as non-specific driving forces for the condensation process, with important implications for the aberrant function, druggability, and material properties of biomolecular condensates. One Sentence SummaryProteins implicated in cellular phase separation can undergo a salt-mediated reentrant liquid-liquid phase transition.

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Section: Introductionsupporting

confidence: 86%

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

Krainer

Welsh

Joseph

et al. 2020

Preprint

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“…Notably, the AFB 1 biodegradation activity of fermentation supernatant of C. funkei was decreased more than 50% after treated with proteinase K or plus SDS, which is similar to the AFB 1 biodegradation by the culture supernatant of F. aurantiacum and S. maltophilia reported earlier, and indicated the active ingredient could therefore be protein or perhaps an enzyme (Alberts et al ., ; Guan et al ., ). Furthermore, the AFB 1 biodegradation activity was positively correlated with the protein content from the fermentation supernatant of C. funkei by ammonium sulfate precipitation (http://onlinelibrary.wiley.com/doi/10.1111/1751-7915.12244/suppinfo) which provides further evidence that the active ingredient could be protein (Callejón et al ., ; Duong‐Ly and Gabelli, ). However, heating may cause denaturation of proteins, strikingly, the AFB 1 biodegradation activity of fermentation supernatant of C. funkei was only slightly affected by heating.…”

Section: Discussionmentioning

confidence: 99%

A novel strain of Cellulosimicrobium funkei can biologically detoxify aflatoxin B₁ in ducklings

Sun

Zhang

Sun

et al. 2015

Microbial Biotechnology

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Two experiments were conducted to screen microorganisms with aflatoxin B1 (AFB1) removal potential from soils and to evaluate their ability in reducing the toxic effects of AFB1 in ducklings. In experiment 1, we screened 11 isolates that showed the AFB1 biodegradation ability, and the one exhibited the highest AFB1 removal ability (97%) was characterized and identified as Cellulosimicrobium funkei (C. funkei). In experiment 2, 80 day-old Cherry Valley ducklings were divided into four groups with four replicates of five birds each and were used in a 2 by 2 factorial trial design, in which the main factors included administration of AFB1 versus solvent and C. funkei versus solvent for 2 weeks. The AFB1 treatment significantly decreased the body weight gain, feed intake and impaired feed conversion ratio. AFB1 also decreased serum albumin and total protein concentration, while it increased activities of alanine aminotransferase and aspartate aminotransferase and liver damage in the ducklings. Supplementation of C. funkei alleviated the adverse effects of AFB1 on growth performance, and provided protective effects on the serum biochemical indicators, and decreased hepatic injury in the ducklings. Conclusively, our results suggest that the novel isolated C. funkei strain could be used to mitigate the negative effects of aflatoxicosis in ducklings.

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“…After ammonium sulfate precipitation (Duong‐Ly and Gabelli, 2014), the vast majority of the DepA activity was found among the proteins that precipitated between 50 and 70% of saturating conditions. This fraction was then dialysed to remove excess ammonium sulfate and heated at 60°C to precipitate less stable proteins.…”

Section: Resultsmentioning

confidence: 99%

The enzymatic detoxification of the mycotoxin deoxynivalenol: identification of DepA from the DON epimerization pathway

Carere

Hassan

Lepp

et al. 2017

Microbial Biotechnology

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SummaryThe biological detoxification of mycotoxins, including deoxynivalenol (DON), represents a very promising approach to address the challenging problem of cereal grain contamination. The recent discovery of Devosia mutans 17‐2‐E‐8 (Devosia spp. 17‐2‐E‐8), a bacterial isolate capable of transforming DON to the non‐toxic stereoisomer 3‐epi‐deoxynivalenol, along with earlier reports of bacterial species capable of oxidizing DON to 3‐keto‐DON, has generated interest in the possible mechanism and enzyme(s) involved. An understanding of these details could pave the way for novel strategies to manage this widely present toxin. It was previously shown that DON epimerization proceeds through a two‐step biocatalysis. Significantly, this report describes the identification of the first enzymatic step in this pathway. The enzyme, a dehydrogenase responsible for the selective oxidation of DON at the C3 position, was shown to readily convert DON to 3‐keto‐DON, a less toxic intermediate in the DON epimerization pathway. Furthermore, this study provides insights into the PQQ dependence of the enzyme. This enzyme may be part of a feasible strategy for DON mitigation within the near future.

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Salting out of Proteins Using Ammonium Sulfate Precipitation

Cited by 230 publications

References 5 publications

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

A novel strain of Cellulosimicrobium funkei can biologically detoxify aflatoxin B₁ in ducklings

The enzymatic detoxification of the mycotoxin deoxynivalenol: identification of DepA from the DON epimerization pathway

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Salting out of Proteins Using Ammonium Sulfate Precipitation

Cited by 230 publications

References 5 publications

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

A novel strain of Cellulosimicrobium funkei can biologically detoxify aflatoxin B1 in ducklings

The enzymatic detoxification of the mycotoxin deoxynivalenol: identification of DepA from the DON epimerization pathway

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A novel strain of Cellulosimicrobium funkei can biologically detoxify aflatoxin B₁ in ducklings