酵母におけるERーゴルジ間輸送の分子機構

酵母内质网-高尔基体转运的分子机制

• 批准号: 01580185
• 负责人: 中野 明彦
• 金额: $ 1.22万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1989
• 资助国家: 日本
• 起止时间: 1989 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01580185/
• 关键词: タンパク質の分秘; 小胞体(ES); ゴルジ体; 小胞輸送; GTP結合タンパク質; SARI; SEC12; 酵母(S.cerevisial); タンパク質の分泌; 小胞体(ER); 細胞内小胞輸送; SAR1; 酵母(S.cerevisiae)

酵母遺伝子SARIは,ERーゴルジ体間輸送に温度感受性の欠陥をもつ sec12変異株のマルチコピ-サプレッサ-として分離されたものであり,その抑圧の機構は産物レベルでの相互作用によるものと推定されている.その産物Sarlpについては、これまでに,分子量21KDのGTP結合タンパク質であること,それ自身がERーゴルジ体間輸送に必須であること,そして細胞内では主に小胞体膜に局在するが可溶性のも存在することを示してきた.本年度は,さらにERーゴルジ体間輸送の無細胞系を用い,以下のことを明らかにした.酵母のin vitro ERーゴルジ体間輸送を,Bakerらの報告に基づき,α因子の糖鎖修飾を指標として確立した.この反応で,sec12変異株より調製したsemiーintact cells(膜画分)は,in vitroでもin vivoと同様に温度感受性を示した.この系にSarlp過剰生産株より調製した細胞質可溶性画分を加えたところ,sec12膜の温度感受性はほぼ完全に抑圧された.この抑圧効果はSarlp自身によるものであることが示されたので,in vivoにおけるSarlpの生理活性をin vitroで再現できたものと考えられる.野生型およびGTP非結合型の変異Sarlpを大腸菌で作らせたところ,野生型Sarlpのみが活性を示した.また,GTPの非分解アナログであるGTPγSを結合した野生型Sarlpも活性をもたなかった.つまり,SarlpのERーゴルジ間輸送の機能には,GTPの結合とその水解が必須であることが示唆された.

The temperature sensitivity of yeast yeast SARI, ER yeast was not sensitive to temperature, and the temperature sensitivity of sec12 strain was not sensitive to temperature. The temperature was isolated from yeast, sec12, yeast, yeast. The molecular weight of the compound, Sarlp, GTP, molecular weight, molecular weight, temperature, temperature, temperature This year, we will send cell-free cell lines to ER parents this year. The following information is available to you. Yeast in vitro ER data transfer system, Baker report data base, alpha factor sugar modification indicates that you must make sure that you are in operation. The temperature sensitivity of sec12 strain is different from that of semi intact cells, and the temperature sensitivity of in vitro is the same as that of temperature sensitivity. The temperature sensitivity of sec12 membrane was completely inhibited by Sarlp through the analysis of cell solubility and temperature sensitivity of SEC 12 membrane. The results show that the Sarlp itself has a negative effect on the physiological activity of the Sarlp, and that the physiological activity of the in vitro has been detected again. The GTP of wild type was used as antifungal agent, and the activity of wild type of Sarlphia spp. GTP was shown by Elisa. The activity of wild-type GTP was detected by the combination of GTP γ S and non-decomposing Sarlp. It is necessary for GTP to transmit the energy of the machine, and it is necessary for Sarlp to combine it with the hydrolysis process.

项目成果

Akihiko Nakano and Masaki Muramatsu: "A Novel GTP-binding Protein,Sar1p,is Involved in Transport from the Endoplasmic Reticulum to the Golgi Apparatus" J.Cell Biol.109. 2677-2691 (1989)

Akihiko Nakano 和 Masaki Muramatsu：“一种新型 GTP 结合蛋白 Sar1p 参与从内质网到高尔基体的运输”J.Cell Biol.109。

T.Oka,S.Nishikawa and A.Nakano: "GTPーdependent reconstitution of the Sorl protein function in vitro:transport from the endoplasmic reticulum to the Golg:apparatus."

T.Oka、S.Nishikawa 和 A.Nakano：“Sorl 蛋白功能的体外 GTP 依赖性重建：从内质网转运至高尔格：装置。”

Akihiko Nakano and Shuh-ichi Nishikawa: "A Novel GTP-binding Protein,Sar1p,Cooperates with Sec12p in Transport from the Endoplasmic Reticulum to the Golgi Appartus" J.Cell Biochem,Suppl.14C. 63 (1990)

Akihiko Nakano 和 Shuh-ichi Nishikawa：“一种新型 GTP 结合蛋白 Sar1p，与 Sec12p 合作从内质网运输到高尔基体”J.Cell Biochem，Suppl.14C。

C.d'Enfert,C.Barlowe,S.Nishikawa,A.Nakano,and R.Schekman: "Functional dissection of Sec12p and evidence for its interaction with a small GTPーbinding protein"

C. dEnfert、C. Barlowe、S. Nishikawa、A. Nakano 和 R. Schekman：“Sec12p 的功能剖析及其与小型 GTP 结合蛋白相互作用的证据”

Shuhーichi Nishikawa and Akihiko Nakano: "The GTPーbinding Sari protein is localized to the early compartment of the yeast secrctory pathway." Biochim.Biophys.Acta.

Shuhiichi Nishikawa 和 Akihiko Nakano：“GTP 结合 Sari 蛋白定位于酵母分泌途径的早期区室。”