Stability of human growth hormone: influence of methionine oxidation on thermal folding.
Oxidation, particularly of methionine residues, is one of the major chemical degradations of proteins. In a previous publication we studied the conformation of recombinant human growth hormone (r-hGH) selectively oxidized at Met14 and Met125. Conformation of oxidized r-hGH was found not different from that of nonoxidized r-hGH. In this paper, the effect of methionine oxidation on the thermal stability of r-hGH folding was investigated. The thermally induced unfolding process of the oxidized and nonoxidized protein was measured by monitoring the circular dichroism signal at 220 nm. The melting temperatures (T(m)) of the oxidized and nonoxidized r-hGH forms were determined at different pHs and in the presence of salts often used in pharmaceutical formulations. The effect of the location of the oxidized Met residue in the protein and the percentage of oxidation were investigated. Our findings indicate that the monoxidation of the two most accessible methionine residues of r-hGH-Met14 and Met125 - has no effect on the protein conformation. However, oxidation of these residues to form sulfoxides does influence the thermal stability of the protein folding. The presence of the polar oxygen atom on the methionine sulfoxide group thermally destabilizes r-hGH folding. The effect (T(m)) depends upon pH, ionic strength, and the location of the oxidized methionine residues in the protein. The thermal melting of r-hGH and its oxidized products is a highly cooperative process. Methionine oxidation leads to a thermal destabilization of the whole protein folding and is not just a local destabilization.
School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Geneva, Switzerland.
Filippo
F
Mulinacci
Martinus A H
MA
Capelle
Robert
R
Gurny
Alex F
AF
Drake
Tudor
T
Arvinte
D013006:Growth Hormone / Q000737:chemistry*; D006801:Humans; D008715:Methionine / Q000737:chemistry*; D010084:Oxidation-Reduction; D055550:Protein Stability; D058767:Protein Unfolding; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature
D011994:Recombinant Proteins; D013006:Growth Hormone; D008715:Methionine