title: Limited proteolysis of disulfide-reduced ovalbumin by subtilisin.
abstract: Our previous study [Takahashi et al., J. Biochem., 109, 846-851 (1991)] has shown that the disulfide-reduced form of ovalbumin was proteolyzed by subtilisin into three major fragments. It was investigated whether or not these three fragments would be folded into one molecule. Gel permeation and ion-exchange chromatography indicated that the three fragments were eluted in a single peak. The proteolyzed protein had a CD spectrum that was almost indistinguishable from the disulfide-reduced, non-proteolyzed form of ovalbumin. Differential scanning calorimetry, however, revealed that the proteolyzed ovalbumin was denatured at a lower temperature than that of the disulfide-reduced, non-proteolyzed protein. Thus, it is concluded that the three fragments were folded into a native-like conformation with decreased stability. Chemical analyses of the fragments purified by reverse-phase HPLC revealed that there was a cleavage site in the disulfide-reduced form of ovalbumin, at least at the amino-terminal side of Cys(73), in addition to the well-known cleavage sites in plakalbumin.
authors.affiliation: Research Institute for Food Science, Kyoto University, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Yun
authors.forename: H
authors.initials: H
authors.lastname: Yamashita
authors.forename: N
authors.initials: N
authors.lastname: Takahashi
authors.forename: M
authors.initials: M
authors.lastname: Hirose
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D004220:Disulfides; D004229:Dithiothreitol; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010047:Ovalbumin / Q000737:chemistry*; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D017510:Protein Folding; D013381:Subtilisins / Q000378:metabolism*; D013696:Temperature
chemical_list: D000596:Amino Acids; D004220:Disulfides; D010446:Peptide Fragments; D010047:Ovalbumin; D013381:Subtilisins; D004229:Dithiothreitol