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Thermodynamic characterization of an intermediate state of human growth hormone
Author(s) -
GomezOrellana Isbuel,
Varinano Bruce,
MiuraFraboni Judy,
Milstein Sam,
Paton Duncan R.
Publication year - 1998
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560070611
Subject(s) - circular dichroism , cooperativity , chemistry , differential scanning calorimetry , crystallography , aqueous solution , protein secondary structure , protein tertiary structure , denaturation (fissile materials) , calorimetry , nuclear chemistry , biochemistry , thermodynamics , physics
The thermal denaturation of recombinant human growth hormone (rhGH) was studied by differential scanning calorimetry and circular dichroism spectroscopy (CD). The thermal unfolding is reversible only below pH 3.5, and under these conditions a single two‐state transition was observed between 0 and 100°C. The magnitudes of the Δ H and Δ C P of this transition indicate that it corresponds to a partial unfolding of rhGH. This is also supported by CD data, which show that significant secondary structure remains after the unfolding. Above pH 3.5 the thermal denaturation is irreversible due to the aggregation of rhGH upon unfolding. This aggregation is prevented in aqueous solutions of alcohols such as n ‐propanol, 2‐propanol, or 1,2‐propanediol (propylene glycol), which suggests that the self‐association of rhGH is caused by hydrophobic interactions. In addition, it was found that the native state of rhGH is stable in relatively high concentrations of propylene glycol (up to 45% v/v at pH 7‐8 or 30% at pH 3) and that under these conditions the thermal unfolding is cooperative and corresponds to a transition from the native state to a partially folded state, as observed at acidic pH in the absence of alcohols. In higher concentrations of propylene glycol, the tertiary structure of rhGH is disrupted and the cooperativity of the unfolding decreases. Moreover, the CD and DSC data indicate that a partially folded intermediate with essentially native secondary structure and disordered tertiary structure becomes significantly populated in 70‐80% propylene glycol.