Role of the Carbohydrate Chain and Two Phosphate Moieties in the Heat-Induced Aggregation of Hen Ovalbumin

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概要

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• We investigated the effect of the carbohydrate chain and two phosphate moieties on heat-induced aggregation of hen ovalbumin. The dephosphorylated form of ovalbumin was obtained by treating the original protein with acid phosphatase. The single carbohydrate chain was removed by digestion of heat-denatured ovalbumin with glycopeptidase F, and the resulting polypeptide without this carbohydrate chain was correctly refolded to acquire protease-resistance. Thermal unfolding can be approximated by a mechanism involving a two-state transition between the folded and unfolded states with a midpoint temperature of 76 °C for the original form, of 74 °C for the dephosphorylated form, and of 71 °C for the carbohydrate-free form. The conformational stability of the original form was higher than that of the carbohydrate-free form. When the three forms of ovalbumin were heated to 80 °C and then cooled rapidly in an ice bath, the polypeptide chains were compactly collapsed to metastable intermediates with secondary structures whose properties were indistinguishable. Upon incubation at 60 °C, renaturation was possible for a large portion of the intermediates of the original form, but for only a small portion of those of the carbohydrate-free form. Light scattering experiments showed that in the presence of sulfate anions, the intermediates of the carbohydrate-free form aggregated to a greater extent than did those of the original form. The intermediates of the carbohydrate-free form bound to the chaperonin GroEL with about 10-fold higher affinity than those of the original form. It follows that the carbohydrate chain and the two phosphate moieties do not affect hydrophobic collapse in the kinetic refolding of hen ovalbumin but play an important role in the slow rearrangement. They block the off-pathway reaction that competes with correct refolding by effectively decreasing surface hydrophobicity.

• 社団法人 日本農芸化学会の論文
• 2004-12-23

著者

• YASUMOTO Kyoden

  Department of Human Nutrition, School of Life Studies, Sugiyama Jogakuen University

• Kitabatake Naofumi

  Research Institute Of Food Science Kyoto University

• Kitabatake Naofumi

  Division Of Food Science And Biotechnology Graduate School Of Agriculture Kyoto University

• Tani Fumito

  Department Of Food Science And Technology Faculty Of Agriculture Kyoto University

• Tani Fumito

  Research Institute For Food Science Kyoto University

• Tani F

  Institute For Fundamental Research Of Organic Chemistry Kyushu University

• Fumito Tani

  Division Of Food Science And Biotechnology Graduate School Of Agriculture Kyoto University

• Tani F

  Institute For Materials Chemistry And Engineering Kyushu University

• Tani Fumito

  Laboratory Of Food And Environmental Sciences Division Of Food Science And Biotechnology Graduate Sc

• Tani Fumito

  Division Of Food Science And Biotechnology Graduate School Of Agriculture Kyoto University

• Tani Fumito

  Institute For Materials Chemistry And Engineering Kyushu University

• Nakanishi Yoko

  Division Of Science Of Biological Resources Graduate School Of Science And Technology Kobe Universit

• SHIRAI Nobuaki

  Industrial Research Center of Shiga Prefecture

• NAKANISHI Yukiko

  Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University

• Nakanishi Y

  Division Of Food Science And Biotechnology Graduate School Of Agriculture Kyoto University

• Kitabatake N

  Division Of Food Science And Biotechnology Graduate School Of Agriculture Kyoto University

• Yasumoto K

  Department Of Food And Nutrition School Of Life Studies Sugiyama Jogakuen University

• Yasumoto Kyoden

  Department Of Food And Nutrition School Of Life Studies Sugiyama Jogakuen University

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