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Characterizing the Specificity of Osmolyte Effects on Protein Thermal Stability
Author(s) -
Canepa Jacob,
Torgerson Julie,
Kim Da Eun,
Lindahl Elizabeth,
Takahashi Rei,
Heying Michael,
Wilkinson Steven
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.05176
Subject(s) - osmolyte , chemistry , amino acid , biochemistry , denaturation (fissile materials) , biophysics , biology , nuclear chemistry
Osmolytes are small organic molecules that demonstrate the ability to shift protein unfolding equilibria when present as solution components, and provide proteins with protection from denaturation against thermal stress. While many subclasses of osmolytes are known, it is unclear whether there is an intrinsic stabilization hierarchy among the molecules that applies to all proteins. Additionally, it is uncertain what physicochemical properties of osmolytes confer their ability to act as stabilizers or destabilizers in solution. Our investigation utilizes differential scanning fluorimetry (DSF) to measure protein stability in two structure‐based surveys: 1) determining whether there is a stabilization hierarchy among the three main chemical classes of osmolytes with respect to the model proteins, human C‐reactive protein (CRP) and tumor necrosis factor alpha (TNFα), and 2) investigating the structural moiety requirements for amino acid osmolytes in the stabilization of CRP and myoglobin. The former part of our investigation reveals that ectoine is the most effective stabilizer of CRP, yet is a destabilizer when acting on TNFα, suggesting that there is no one osmolyte, or class of osmolytes, that confers the greatest stability across multiple proteins. The latter part of our investigation reveals that myoglobin is stabilized only by zwitterionic amino acids, as removal of either the carboxylate or amino moiety from the amino acid abrogates all thermal stabilizing effects. In contrast, in the case of CRP the carboxylate proved to be the dominant stabilizing group, as compounds lacking the amino group conferred slightly greater thermal stability than the complete amino acid osmolyte. Collectively, our results affirm that osmolyte effects on protein thermal stability are highly protein‐specific. In addition, our data suggests that contemporary models to explain the osmolyte effect may not apply to all protein systems. These results may have significant implications for the development of chemistries aiming to stabilize diverse protein targets in biospecimens for diagnostic applications.