Structural properties of recombinant ovalbumin and its transformation into a thermostabilized form by alkaline treatment.
The recombinant ovalbumin produced in Escherichia coli was purified from the cytoplasmic fraction and analyzed for its chemical and conformational properties. The recombinant ovalbumin displayed almost exactly the same circular dichroism and intrinsic tryptophan fluorescence spectra as egg white ovalbumin. As in the egg white protein, four cysteine sulfhydryls and one cystine disulfide were contained in the recombinant protein, according to the results of amino acid analyses; the disulfide bond was found by a peptide mapping analysis to correspond to the native cystine, Cys73-Cys120. According to a gel electrophoresis analysis, the presence of the disulfide bond was accounted for by specific oxidation of the corresponding cysteine residues during purification of the cytoplasmic protein. Unlike the identity in the conformational and peptide structures, none of the post-translational modifications (N-terminal acetylation, phosphorylation, and glycosylation) that are known with egg white ovalbumin were detected in the recombinant protein. The recombinant ovalbumin was transformed into a thermostabilized form in a similar manner to the transformation of egg white protein into S-ovalbumin; alkaline treatment increased the temperature for thermostability by 8.7 degrees C. These data strongly suggest that the post-translational modifications of ovalbumin are not related to the formation mechanism for S-ovalbumin.
Arii
Y
Y
Research Institute for Food Science, Kyoto University, Japan.
Takahashi
N
N
Tatsumi
E
E
Hirose
M
M
D000468:Alkalies / Q000737:chemistry*; D002942:Circular Dichroism; D003593:Cytoplasm / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D010047:Ovalbumin / Q000737:chemistry*; D010449:Peptide Mapping; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013696:Temperature
D000468:Alkalies; D004220:Disulfides; D011994:Recombinant Proteins; D010047:Ovalbumin