ᮏ❶䛷䛿䠈䝅䝱䝨䝻䝙䞁
GroEL
䜘䜚༢㞳䛧䛯䜰䝢䜹䝹䝗䝯䜲䞁(GroEL-AD) 䜢స〇䛧䠈A䃑1-42䠈䃐-Synuclein䠈GroES
䛾䜰䝭䝻䜲䝗⥺⥔䛻ᑐ䛩䜛จ㞟ᢚไຠᯝ䜢᳨ウ䛧䛯䚹㐣㔞䛾GroEL-AD
䜢ῧຍ䛧䛯᮲௳䛻䛚䛔䛶A䃑1-42䠈䃐-Synuclein䠈GroES
䛾䜰䝭䝻䜲䝗⥺⥔ᙧ ᡂ䛿䛻ᢚไ䛥䜜䜛䛾䜢♧䛧䛯(ᅗ4-4)䚹䃐-Synuclein
䜔GroES
䛾⥺⥔ᙧᡂ䛻䛚䛔䛶䛿䠈 㐣㔞䛾GroEL-AD
䜢ῧຍ䛧䛺䛟䛶䜒䠈༑ศ䛺ᢚไຠᯝ䛜䜏䜙䜜䛯䚹䛧䛛䛧䠈A䃑1-42
䛷䛿䛺ᢚไຠᯝ䜢ᚓ䜛䛯䜑䛻䛿
20
ಸ㔞௨ୖ䛾GroEL-AD
䛜ᚲせ䛷䛒䛳䛯䚹䃐-Synuclein 䜔GroES
䛷䛿䛺ᢚไ䛜䜏䜙䜜䛯⃰ᗘ䛻䛚䛔䛶䜒䠈A䃑1-42 䛷䛿䛺ᢚไ䜢♧䛥䛺䛛 䛳䛯(ᅗ4-4d)䚹䛻⥺⥔䜢ᢚไ䛩䜛㔞䜘䜚ప䛔⃰ᗘ䛾 GroEL-AD
䜢ῧຍ䛩䜛䛸䠈จ㞟 ഴྥ䛾䛒䜛䝍䞁䝟䜽㉁ศᏊ䛸┦స⏝䛩䜛䛣䛸䛷䠈᭱⤊ⓗ䛻ᙧᡂ䛩䜛⥺⥔㔞䛜ῶᑡ䛧䛶䛔䜛䛸⪃䛘䜙䜜䛯䚹GroEL-AD 䛾Ꮡᅾୗ䛻䛚䛔䛶ᙧᡂ䛧䛯⥺⥔䛾ᙧែ䛻䛿㐪䛔䛜䜏䜙䜜䛺䛛䛳䛯䚹 䛣䜜䛿䠈ᚑ᮶⪃䛘䜙䜜䛶䛔䜛䜰䝭䝻䜲䝗⥺⥔䜢ᙧᡂ䛩䜛ᶵᵓ䛜ᫎ䛥䜜䛶䛔䜛䛸⪃䛘䜙䜜䜛(ᅗ
4-5䠈ᅗ 4-6)䚹䛧䛛䛧䠈䃐-Synuclein
䛾ሙྜ䛿እ䛷䠈GroEL-ADᏑᅾୗ䛻䛚䛔䛶ᙧᡂ䛧䛯⥺⥔䛾
TEM
⏬ീ䛿䃐-Synuclein ༢⊂ཬ䜃⥺⥔ᙧᡂ䛻↓㛵ಀ䛺BSA
䛾Ꮡᅾୗ䛻䛚䛔䛶 ᙧᡂ䛧䛯⥺⥔䜘䜚䜒⣽䛟䜏䜙䜜䛯(ᅗ4-6)䚹ᅗ 4-9
䛾QCM
ᐇ㦂䛻䛚䛡䜛GroEL-AD 䛸䃐-Synuclein
㛫 䛾 ┦ స ⏝ ほ 䛥 䜜 䛯 」 ᩘ 䛾 ⤖ ྜ ᵝ ᘧ 䛾 䛯 䜑 䛻 䠈GroEL-AD
䛜 䃐-Synuclein
䛾⥺⥔ఙ㛗䜢ᢚไ䛧䛶䛚䜚䠈䛥䜙䛻ᙧᡂ䛩䜛⥺⥔䛾ᙧែ䜢ኚ䛩䜛⬟ຊ䜒᭷䛧䛶䛔䜛䛣䛸䛜⪃䛘䜙䜜䛯䚹
ᮏ❶䛻䛚䛔䛶☜ㄆ䛥䜜䛯GroEL-AD䛾ᵝ䚻䛺ᶆⓗ䝍䞁䝟䜽㉁䛾⥺⥔ᙧᡂ䜢ᙉᅛ䛻ᢚไ䛩 䜛⬟ຊ䛿䠈ᵝ䚻䛺ᐇ㦂᮲௳ୗ䛻䛚䛔䛶䛥䜙䛻᫂䜙䛛䛸䛺䛳䛯䚹䜎䛪䠈GroEL-AD䛿䠈42䜰䝭䝜 㓟 ṧ ᇶ 䛾 ▷ 䛔A䃑
1-42
䝨 䝥䝏 䝗 䛸 䠈97䜰䝭 䝜 㓟 ṧ ᇶ 䛾GroES
ཬ 䜃140䜰䝭 䝜 㓟 ṧ ᇶ 䛾䃐-Synuclein
䛾ᬑ㏻䛾䛝䛥䛾୧᪉䜢ᶆⓗ䛸䛧䛶⤖ྜ䛷䛝䛯䚹䛭䜜䛻ຍ䛘䠈䛣䜜䜙ᶆⓗ䛸䛺䜛䝍 䞁 䝟 䜽 ㉁ 䛾 ᵓ 㐀 䛿 ᵝ 䚻 䛷 䠈 ▷ 䛔 䝫 䝸 䝨 䝥䝏 䝗䛷 䛒 䜛A
䃑1-42䠈 ኳ ↛ ኚ ᛶ 䝍䞁 䝟 䜽 ㉁ 䛾 䃐-Synuclein䠈䛭䛧䛶䝛䜲䝔䜱䝤ᵓ㐀䛾䜸䝸䝂䝬䞊䛜㒊ศⓗ䛻ኚᛶ䛧䛯 GroES䛜Ꮡᅾ䛩䜛䚹䛭䜜
121
䛻䜒䛛䛛䜟䜙䛪䠈ಶ䚻䛾䝍䞁䝟䜽㉁䛻䛚䛔䛶ほ 䛥䜜䜛䛭䜜䛮䜜䛾⥺⥔ ᙧ ᡂ ᮲ ௳䛻䛚䛔䛶
GroEL-AD䛿⤖ྜ䛻క䛔䜰䝭䝻䜲䝗⥺⥔จ㞟䜢ไᚚ䛩䜛䛣䛸䛜䛷䛝䛯䚹䛧䛛䛧䠈QCM䛻䜘䜛⤖
ྜ䛾ศᯒ䛻䛚䛔䛶䠈ᶆⓗ 䝍䞁䝟䜽㉁䛸GroEL-AD䛾⤖ྜ䛻䛚䛡䜛⤖ྜᵝᘧ䛿୍ᐃ䛷䛿䛺䛔 䛾䛜☜ㄆ䛥䜜䛯䚹䛣䜜䛿䠈3䛴䛾ᶆⓗ䝍䞁䝟䜽㉁䛸GroEL-AD䛸䛾⤖ྜᶵᵓ䛻䛚䛡䜛␗䛺䜛จ 㞟ᶵᵓ䛜ᫎ䛥䜜䛶䛔䜛䛸⪃䛘䜙䜜䜛䚹
GroEL䛿䠈⣽⬊ෆ䝇䝖䝺䝇䛻䜘䜚୍ⓗ䛻✚䛧䛯䜰䞁䝣䜷䞊䝹䝗䝍䞁䝟䜽㉁ศᏊ䜈䛾Ỉ
ⓗឤཷᛶ䛜䜘䛟▱䜙䜜䛶䛔䜛䚹GroEL䛾䜰䝢䜹䝹䝗䝯䜲䞁䛿䠈䛣䜜䜙䛾ศᏊ䜢༊ู䛧䠈⤖ྜ䛩䜛
Ỉᛶ䝉䞁䝃䞊䛸䛧䛶ാ䛟䝗䝯䜲䞁䛷䛒䜛䛾䛜❧ド䛥䜜䛶䛔䜛(ᅗ4-1䠈䝦䝸䝑䜽䝇H䠈I)䚹ᮏ❶䛷 䛿䛥䜙䛻䠈3䛴䛾䜰䝭䝻䜲䝗⥺⥔䛜⏕䛨䜛ᶆⓗ䝍䞁䝟䜽㉁䛻ᑐ䛩䜛Ỉᛶຠᯝ䜈䛾ᐤ䜢♧
䛧䛯䚹⥺⥔䝍䞁䝟䜽㉁䛻䛚䛡䜛Ỉᛶ䛾ᙺ䛿䠈BSA䜢䝁䞁䝖䝻䞊䝹䛸䛧䛯ᐇ㦂䛻䛚䛔䛶䜘䜚 ヲ⣽䛻☜ㄆ䛥䜜䛯(ᅗ4-6)䚹BSA䛿䠈ᶆⓗ䝍䞁䝟䜽㉁䛻ᑐ䛧䛶ᙳ㡪䛜䛺䛔⬡㉁䛸⤖ྜ䛧䠈⾑ὶ 䜢㏻ 䛧䛶ᵝ 䚻䛺ᰤ 㣴 䛸䛺䜛ศ Ꮚ 䜢㍺ ㏦ 䛩䜛⾑ Ύ 䝍䞁䝟䜽㉁ 䛷䛒䜛䚹BSA䛿䠈A䃑1-42
䠈䃐-Synuclein䠈GroES
䛾⥺ ⥔ ᙧ ᡂ 䛻䛚䛔䛶ᙳ 㡪 䜢ཬ 䜌䛧䛶䛔䜛䜘䛖䛻䜏䜙䜜䜛䛜䠈➼ ⃰ ᗘ 䛾GroEL-AD䜢ῧຍ䛧䛯᮲௳䛸ẚ㍑䛧䛶䛭䛾ᙳ㡪䛿ᙅ䛔(ᅗ4-6)䚹䛣䛾䝁䞁䝖䝻䞊䝹ᐇ㦂䛻䛚䛔
䛶䠈GroES䛷䛿᰾ᙧᡂ䛻䠈A䃑1-42䛷䛿ᙧᡂ䛥䜜䜛⥺⥔䛾ᙧែ䛜ⴭ䛧䛟ኚ䛧䛶䛔䜛䜘䛖䛻䠈BSA䛿䛭䜜䛮䜜䛾ᶆⓗ䝍䞁䝟䜽㉁䛾⥺⥔ᙧᡂ䛻ᑐ䛧␗䛺䜛ᙳ㡪䜢ཬ䜌䛧䛶䛔䜛䚹䛣䛾⤖ᯝ䜘
䜚䠈䜰䝭䝻䜲䝗⥺⥔ᙧᡂ㐣⛬䛾᰾ᙧᡂ䛸⥺⥔ఙ㛗䛻䛚䛡䜛Ỉᛶ┦స⏝䛿ከ✀ከᵝ䛺ᵝᘧ 䜢䜒䛳䛶䛔䜛䛣䛸䛜♧䛥䜜䛯䚹䛭䜜䛻䠈A䃑1-40䛾⥺ ⥔ᙧ ᡂ䛻㛵䛩䜛௨ ๓ 䛾◊✲䛻䛚䛔䛶䠈 ᵝ䚻䛺ṇ䛸㈇䛾㟁Ⲵ䜢䜒䛴ྜ≀䜢⏝䛔䛯⥺⥔䛻䛚䛡䜛㟼㟁┦స⏝䛜㛵䛧䛶䛔䜛䛣䛸 䜒ሗ ࿌ 䛥䜜䛶䛚䜚䠈 Ỉ ᛶ ┦ స ⏝ 䛰䛡䛜⥺ ⥔ 䜢ㄪ ᩚ 䛧䛶䛔䜛䛾䛷䛿䛺䛔䛸⪃ 䛘䜙䜜䜛(115,116)䚹እᅉ䛻䜘䜛่⃭䜈䛾ឤཷᛶ䛻ᑐ䛩䜛䝍䞁䝟䜽㉁䛾⥺⥔䛻䛿ከ䛟䛾┦స⏝䛜
㛵䛧䛶䛔䜛䚹䛣䜜䛿䠈䝍䞁䝟䜽㉁䛸䝍䞁䝟䜽㉁䛾┦స⏝䛻䜘䜛⥺⥔䛾NMR䜔㉁㔞ศᯒ 䛺䛹䛾ཎᏊ䝺䝧䝹䛻䛚䛡䜛ศᯒ䜘䜚᫂䜙䛛䛸䛺䜛䛷䛒䜝䛖䚹122
➨
➨ 5 ❶㻌 ⥲ྜ⪃ᐹ
ᮏ◊✲䛷䛿䠈➨
2
❶䛷GroEL G192X
ኚ␗య䛻䜘䜛䝅䝱䝨䝻䝙䞁ᶵ⬟ゎᯒ䠈➨3
❶䛷GroEL G192X
䜢⏝䛔䛯ኳ↛ኚᛶ䝍䞁䝟䜽㉁䛾┦స⏝䛸䜰䝭䝻䜲䝗⥺⥔ᙧᡂ䜈䛾㛵䛻䛴䛔䛶䠈➨
4
❶䛷䛿GroEL
䛾䜰䝢䜹䝹䝗䝯䜲䞁䜢༢㞳䛧䛯GroEL-AD
䜢⏝䛔䛶䠈䜰䝭䝻䜲䝗⥺⥔ᙧᡂ䜈䛾㛵䜢ホ౯䛧䛯䚹ᮏ❶䛷䛿௨ୖ䛾⤖ᯝ䜢䜅䜎䛘䛶䠈GroEL 䛻䛚䛡䜛䜰䝢䜹䝹䝗䝯䜲 䞁䛜㛵䛧䛶䛔䜛䜰䝭䝻䜲䝗⥺⥔ᙧᡂ䜈䛾㔜せ䛺ᙺ䛻䛴䛔䛶⥲ྜ⪃ᐹ䛩䜛䚹
➨
2
❶䛻䛚䛔䛶GroEL WT
䛾Hinge䊡㒊䛻䛚䛡䜛 Gly192
ṧᇶ䜢䝣䜯䞁䝕䝹䝽䞊䝹䝇 య✚䛻ὀ┠䛧䛶ᵝ䚻䛺䜰䝭䝜㓟ṧᇶ䛻⨨䛧䛯GroEL G192X
ኚ␗య䛿䠈䛝䛔䜰䝭䝜㓟ṧ ᇶ䛻⨨䛧䛯ኚ␗య䜋䛹䝅䝱䝨䝻䝙䞁ᶵ⬟䛻ᙳ㡪䜢ཬ䜌䛧䛶䛔䜛䛣䛸䜢♧䛧䛯䚹䛭䛣䛷䠈5-1 䛻 䛶GroEL
䛾Hinge
䊡㒊䛻䛚䛡䜛Gly192
ṧᇶ䛾ᙺ䛻䛴䛔䛶㏙䜉䜛䚹䛥䜙䛻䠈➨
3
❶䜘䜚GroEL G192X
ኚ␗య䛿ኚ␗య㛫䛻䛚䛔䛶⥺⥔ᙧᡂ䛾␗䛺䜛㐣⛬䛻 ᙳ㡪䜢ཬ䜌䛧䠈ᙧᡂ䛥䜜䛯⥺⥔䛾ᙧែ䜒ኚ䛧䛶䛔䛯䚹䛭䛣䛷䠈5-2 䛻䛶GroEL
䛾Gly192
ṧᇶ䛾ኚ䛧䛯䛣䛸䛻䜘䜛⥺⥔ᙧᡂ䜈䛾ᙳ㡪䛻䛴䛔䛶⪃ᐹ䛩䜛䚹䜎䛯䠈➨
4
❶䜘䜚䠈GroEL-AD䛜䜰䝭䝻䜲䝗⥺⥔ᙧᡂ䛾ᢚไ䜢⾜䛳䛶䛔䜛䛾䜢♧䛧䛯䚹䛭䛣䛷䠈5-3
䛻䛶䜰䝭䝻䜲䝗⥺⥔䜢ᙧᡂ䛩䜛䝍䞁䝟䜽㉁䛻ᑐ䛩䜛GroEL-AD
䛾ᐤ䛻䛴䛔䛶㆟ㄽ䛩䜛䚹5-1䠊Hinge䊡㒊䛻Ꮡᅾ䛩䜛 Gly192 ṧᇶ䛾 GroEL 䛻䛚䛡䜛ᙺ
GroEL
䛾Hinge䊡㒊䛻䛚䛡䜛 Gly192
ṧᇶ䛿䠈䜰䝭䝜㓟ṧᇶ䜢ኚ䛥䛫䛶䜒䛭䛾ḟ ᵓ㐀䛸ᅄḟᵓ㐀䛻䛝䛺㐪䛔䛿䛺䛛䛳䛯(ᅗ2-5䠈2-7)䚹୍᪉䛷䠈ANS
⺯ග ᐃ䛻䜘䜛⾲㠃Ỉᛶ䛾ゎᯒ䛛䜙䠈Gly192ṧᇶ䛾⨨䛧䛯䜰䝭䝜㓟ṧᇶ䛜䛝䛔䜋䛹⾲㠃Ỉᛶ䛜㧗䛟䛺䛳䛶 䛔䛯(ᅗ
2-4)䚹GroEL
䛾ෆ㒊䛿䠈Ỉᛶ㡿ᇦ䛜ከ䛔䛸䛥䜜䛶䛔䜛(16)䚹Gly192ṧᇶ䜢ኚ䛥 䛫䛯䛣䛸䛷GroEL
䛜Open
ᵓ㐀䛻䛺䜚䠈ANS 䛻䜘䜛GroEL
䛾䜰䝢䜹䝹䝗䝯䜲䞁䜈䛾⤖ྜ䛜 ኚ 䛩 䜛 䛣 䛸 䛷 䠈ANS
⺯ ග ᙉ ᗘ 䛾 ኚ 䛻 ᙳ 㡪 䛧 䛶 䛔 䜛 䛣 䛸 䛜 ⪃ 䛘 䜙 䜜 䛯 䚹Pyrococcus
123
furiosus
⏤᮶䛾䜾䝹䞊䝥䊡ᆺ䝅䝱䝨䝻䝙䞁(PfCPN)䛻䛚䛔䛶䠈Gly345 ṧᇶ䜢Asp
䛻⨨䛧䛯
G345D
ኚ␗య䛿䠈Co(2+)䛾Ꮡᅾୗ䛻䛚䛔䛶㔝⏕ᆺ䛾⣙25%䛧䛛 ATP/ADP
ຍỈศ ゎάᛶ䜢♧䛥䛪䠈in vitro
䛻䛚䛔䛶ኚᛶMDH
䛾䝣䜷䞊䝹䝕䜱䞁䜾ຓ䛾⬟ຊ䛜ῶᑡ䛧䛶䛔 䛯(117)䚹Gly-345 䛿ྂ⣽⳦䜾䝹䞊䝥䊡ᆺ䝅䝱䝨䝻䝙䞁䛻㧗䛟ಖᏑ䛥䜜䛶䛚䜚䠈䜰䝢䜹䝹䝗䝯䜲 䞁䛸䜲䞁䝍䞊䝯䝕䜱䜶䜲䝖䝗䝯䜲䞁䛾㛫㏆䛻Ꮡᅾ䛧䛶䛔䜛䚹䝅䝱䝨䝻䝙䞁䛿ኚᛶᇶ㉁䛾䝸䝣䜷䞊 䝹䝕䜱䞁䜾䜢⿵ຓ䛩䜛㝿䛻つᶍ䛺ᵓ㐀ኚ䛜⏕䛨䜛䛣䛸䛜▱䜙䜜䛶䛔䜛䚹䛭䛾䛯䜑䠈䝅䝱䝨 䝻䝙䞁䛜ᶵ⬟䜢ᯝ䛯䛩䛻䛒䛯䜚䠈ᵓ㐀ኚ䛾ᨭⅬ䛸䛺䜛㒊䛻Ꮡᅾ䛩䜛䜰䝭䝜㓟䜢⨨䛩䜛䛣 䛸䛷䠈ᵓ㐀ኚ䛸䛭䜜䛻క䛖ᶵ⬟Ⓨ⌧䛻䛚䛡䜛ᙺ䜢ヲ⣽䛻ㄪ䜉䜛䛣䛸䛜䛷䛝䜛䚹GroEL 䛻 䛚䛡䜛Gly192
ṧᇶ䜢ኚ䛥䛫䛯䛣䛸䛻䜘䜛䝅䝱䝨䝻䝙䞁ᶵ⬟䜈䛾ᙳ㡪䜢ㄪ䜉䛯䛸䛣䜝䠈ATP 䛾ຍỈศゎ⬟ຊ䠈MDH 䛸Rhodanese
䜢ᇶ㉁䛸䛧䛶⏝䛔䛯䝸䝣䜷䞊䝹䝕䜱䞁䜾ຓ⬟ຊ䛿䠈Gly192
ṧᇶ䜢⨨䛧䛯䜰䝭䝜㓟ṧᇶ䛜䛝䛔䜋䛹䛭䜜䛮䜜䛾άᛶ䛜పୗ䛧䛶䛔䛯䚹ZacharyW䠊White
䜙䛿䠈GroEL 䛻䛚䛡䜛3
␒┠䛾Lys
䜢Glu
ṧᇶ䛻⨨䛧䛯GroEL K3E
ኚ␗య䜢⏝䛔䛯◊✲䛻䛚䛔䛶䠈GroEL K3E 䛿䠈༢㔞యᵓ㐀䛷䛒䜛䛜䠈䛭䛾ḟᵓ㐀䛿
GroEL WT
䛸㢮ఝ䛧䛶䛚䜚䠈GroEL WT䛸ẚ㍑䛧䠈㧗䛔ANS
⺯ග䛜ほ 䛥䜜䛯䚹䛧䛛䛧䠈䝚䜽䝺䜸䝏 䝗Ꮡᅾୗ䛻䛚䛔䛶ኚᛶᇶ㉁䛾䝸䝣䜷䞊䝹䝕䜱䞁䜾䜢⿵ຓ䛩䜛䛣䛸䛜䛷䛝䛺䛛䛳䛯(118)䚹௨๓䠈Gly192
ṧᇶ䜢᭱䜒䛝䛔䜰䝭䝜㓟ṧᇶ䛷䛒䜛Trp
䛻⨨䛧䛯GroEL G192W
ኚ␗య䛿䠈ATP
䛾㠀Ꮡᅾୗ䛻䛚䛔䛶cis䉄 ring
ഃ䛻GroES
䛜⤖ྜ䛩䜛䛣䛸䜢ሗ࿌䛧䛯(19)䚹GroELG192X
ኚ␗య䛾䝅䝱䝨䝻䝙䞁ᶵ⬟ゎᯒ䛾⤖ᯝ䜘䜚䠈GroEL G192Xኚ␗య䛿䠈Gly192ṧᇶ 䜢⨨䛧䛯䜰䝭䝜㓟䛻䜘䜚GroES
䛸䛾┦స⏝䛻ኚ䛜⏕䛨䛶䛔䜛䛣䛸䛜⪃䛘䜙䜜䛯䚹QCM ἲ䛻䜘䜛ATP
㠀Ꮡᅾୗ䛻䛚䛡䜛GroEL G192X
ኚ␗య䛸GroES
䛾┦స⏝䜢ゎᯒ䛧䛯⤖ᯝ䛷䛿䠈Gly192 䜢䛝䛔䜰䝭䝜㓟ṧᇶ䛻⨨䛧䛯ኚ␗య䜋䛹
GroES
䛸䛾┦స⏝䛜ᙉ䛔䛣 䛸䛜᫂䜙䛛䛸䛺䛳䛯(ᅗ2-11)䚹
GroEL G192X
ኚ␗య䛻䛚䛡䜛cis
䠈trans
䉄ring䛾ാ䛝䜢䜘䜚ヲ⣽䛻ゎᯒ䛩䜛䛣䛸䜢┠ⓗ䛸 䛧䠈cis
䉄ring䛾ᶵ⬟䛰䛡䜢䜒䛴GroEL
䛾䝅䞁䜾䝹䝸䞁䜾ኚ␗యSR-1
䛾Gly192
䛻䛚䛡䜛䜰䝭124
䝜㓟ṧᇶ䜢⨨䛧䛯
SR-1 G192X
ኚ␗య䜢⏝䛔䛯䝅䝱䝨䝻䝙䞁ᶵ⬟ゎᯒ䜢⾜䛳䛯䚹MDH 䛾 䝸䝣䜷䞊䝹䝕䜱䞁䜾άᛶ ᐃ䛾⤖ᯝ䛻䛚䛔䛶䠈䝅䞁䜾䝹䝍䞊䞁䜸䞊䝞䞊(94)䛜ཎᅉ䛷άᛶ䜢 䜋䛸䜣䛹♧䛥䛺䛔SR-1 WT
䛸ẚ㍑䛧䛶SR-1 G192V
䛸SR-1 G192I
ኚ␗య䛷䛿㧗䛔άᛶ䜢♧䛧䛶䛔䛯䚹௨๓䛾◊✲䛻䛚䛔䛶䠈䝍䞁䝕䝮䛻⧅䛜䛳䛯
GroES 7
㔞య䛻䛚䛡䜛䛔䛟䛴䛛䛾GroES
䝃䝤䝴䝙䝑䝖䜢Asp
ṧᇶ䛻⨨䛧䛯䛣䛸䛷䠈SR-1
䛸䛾┦స⏝䛿ᙅ䜎䜛䛜䠈ኚᛶMDH
䛾䝸䝣䜷䞊䝹䝕䜱䞁䜾άᛶ䛜GroES WT
䜘䜚䜒ቑຍ䛧䛶䛔䜛䛣䛸䛜ሗ࿌䛥䜜䛶䛔䜛(119)䚹 䛥䜙䛻䠈GroEL 䛸┦స⏝䛩䜛䛣䛸䛜䛷䛝䜛䝃䝤䝴䝙䝑䝖䛾ᩘ䜢ኚ䛥䛫䛯GroES
䛻䛚䛔䛶䠈7
ဨ⎔GroES
䝃䝤䝴䝙䝑䝖ᙜ䛯䜚GroEL
䛸┦స⏝䛩䜛䛣䛸䛜䛷䛝䜛䝃䝤䝴䝙䝑䝖䛜5
䛴Ꮡ ᅾ䛧䛶䛔䜛䛸GroEL WT
䛸GroES WT
䛸ྠ⛬ᗘ䛾䝸䝣䜷䞊䝹䝕䜱䞁䜾άᛶ䜢♧䛩䛣䛸䛜ሗ࿌䛥䜜䛶䛔䜛(120)䚹䛣䛾ሗ࿌䛷䛿䛥䜙䛻
GroES
䛾䝃䝤䝴䝙䝑䝖䛜㞄᥋䛧䛶䛔䜛䜋䛹㔝⏕ᆺ䛻㢮 ఝ䛧䛯ᶵ⬟䜢ಖᣢ䛩䜛䛣䛸䛜♧䛥䜜䛶䛔䜛䚹䛣䜜䜙䛾ሗ࿌䛛䜙䠈ᇶ㉁䛾䝸䝣䜷䞊䝹䝕䜱䞁䜾䜢ຓ䛩䜛䛻䛒䛯䜚䠈GroEL䛸
GroES
䛿7
ဨ⎔ྠኈ䛜䛶⤖ྜ䛧䛶䛔䛺䛔ྍ⬟ᛶ䜒⪃䛘䜙䜜䜛䚹SR-1 G192X
ኚ␗య䛾䝅䝱䝨䝻䝙䞁ᶵ⬟ゎᯒ䛾⤖ᯝ䜘䜚䠈GroEL G192Xኚ␗య䛾cis
䉄ring 䛻䛚䛡䜛GroES
䛸䛾┦స⏝䛿Gly192
ṧᇶ䜢᭱䜒䛝䛔Trp
䛻⨨䛩䜛䛣䛸䛷䛻GroES
䛸⤖ྜ䛩䜛䛜䠈䛭䜜䜘䜚ᑠ䛥䛔䜰䝭䝜㓟䛷䛿䛻䛿GroES
䛸⤖ྜ䛧䛶䛔䛺䛔䛣䛸䛜⪃䛘䜙䜜䜛䚹䛣䜜䜘䜚䠈GroEL G192Xኚ␗య䛻䛚䛡䜛
GroES
䛸䛾┦స⏝䛿䠈Gly192ṧᇶ䜢⨨䛩䜛䜰䝭䝜㓟ṧᇶ䜢㑅ᢥ䛩䜛䛣䛸䛷
Open
≧ែ䛾䜰䝢䜹䝹䝗䝯䜲䞁䜢ㄪ⠇䛩䜛䛣䛸䛷ኚ䛥 䛫䜛䛣䛸䛜ྍ⬟䛻䛺䜛䛸⪃䛘䜙䜜䜛䚹䛥䜙䛻䠈GroES䛸䛾┦స⏝䜢ኚ䛥䛫䜛䛣䛸䛷ኚᛶᇶ㉁䛾䝸䝣䜷䞊䝹䝕䜱䞁䜾ຓ⬟ຊ䜒ㄪ⠇䛩䜛䛣䛸䛜ྍ⬟䛸䛺䜛䛸⪃䛘䜙䜜䛯䚹
௨ୖ䜘䜚䠈Gly192 ṧᇶ䛿䠈GroEL 䛜ᮏ᮶᭷䛧䛶䛔䜛ኚᛶ䝍䞁䝟䜽㉁䛾䝸䝣䜷䞊䝹䝕䜱䞁䜾
⿵ຓ⬟ຊ䜢Ỵᐃ䛩䜛㔜せ䛺ᅉᏊ䛷䛒䜚䠈䛣䛾
Hinge䊡㒊䛻⨨䛩䜛ṧᇶ䜢ኚ䛥䛫䜛䛣䛸
䛷
GroEL
䛾䝅䝱䝨䝻䝙䞁ᶵ⬟䜢䝍䞁䝟䜽㉁ᕤᏛⓗ䛻ㄪᩚ䛩䜛䛣䛸䛜ྍ⬟䛷䛒䜛䛣䛸䛜♧၀䛥䜜䛯䚹䛣䛾䛸䛝䛾⪃䛘䜙䜜䜛䝅䝱䝨䝻䝙䞁䝃䜲䜽䝹䜢ᅗ
5-1
䛻♧䛩䚹125
ᅗ5-1. GroEL WT 䛸GroEL G192Xኚ␗య䛾䝅䝱䝨䝻䝙䞁䝃䜲䜽䝹䛾ẚ㍑
ୖᅗ䠖GroEL G192Xኚ␗య䛾䝅䝱䝨䝻䝙䞁䝃䜲䜽䝹
Hinge䊡㒊䛾Gly192ṧᇶ䜢⨨䛩䜛䛣䛸䛷䜰䝢䜹䝹䝗䝯䜲䞁䛾ഴ䛝䜢ኚ䛥䛫䛶䛔䜛(䕔ෆ)䚹 䛣䛾ഴ䛝䛾ኚ䛿Gly192䜢䝣䜯䞁䝕䝹䝽䞊䝹䝇య✚䛻䜘䜚⨨䛧䛯ṧᇶ䛻䜘䜚ኚ䛩䜛䚹
GroEL G192X ኚ␗య䛿䠈䜰䝢䜹䝹䛜ୖ䛜䛳䛶䛔䜛 cis䉄ring 䛻䛿 ATP㠀Ꮡᅾୗ䛷GroES䛜⤖ྜ
䛩䜛䚹trans䉄ring䛻䛿ኚᛶᇶ㉁䛜⤖ྜ䛩䜛䚹䛭䛾ᚋ䠈ATP䛸䛥䜙䛺䜛GroES䛾ῧຍ䛷trans䉄ring䛾 ኚᛶᇶ㉁䛸 GroES 䛜⨨䛝䜟䜛䚹⨨䛧䛯䜰䝭䝜㓟ṧᇶ䛻䜘䛳䛶䛿୍㒊䛜䜻䝱䝡䝔䜱ෆ䛻ධ䜚㎸䜣䛷䝸 䝣䜷䞊䝹䝕䜱䞁䜾䛾ຓ䛜⾜䜟䜜䜛䚹
ୗᅗ䠖GroEL WT 䛾䝅䝱䝨䝻䝙䞁䝃䜲䜽䝹
GroEL WT 䛿ኚᛶᇶ㉁䜢cis䉄ring䛻⤖ྜ䛥䛫䛯ᚋ ATP䛾Ꮡᅾୗ䛷 GroES䛜⤖ྜ䛩䜛䛣䛸䛷䠈
つᶍ䛺ᵓ㐀ኚ䜢ᘬ䛝㉳䛣䛧ෆ㒊䛻䝉䞁䝖䝷䝹䜻䝱䝡䝔䜱䛸䜀䜜䜛✵Ὕ䜢స䜚ฟ䛧䠈ኚᛶᇶ㉁䜢ⴠ䛸䛧
㎸䜐䚹䛭䛾ᚋኚᛶᇶ㉁䛜Ꮡᅾ䛧䛶䛔䜛≧ែ䛷䛿ኚᛶᇶ㉁䜢 trans䉄ring ഃ䛻⤖ྜ䛩䜛䚹⤖ྜ䛧䛶䛔䜛 ATP䛜ADP䛻ຍỈศゎ䛥䜜䜛䛸GroES䛜እ䜜䠈ኚᛶᇶ㉁䛜እ䛻ᨺฟ䛥䜜䜛䚹
126
55-2䠊ኳ↛ኚᛶ䝍䞁䝟䜽㉁䛾䜰䝭䝻䜲䝗⥺⥔ᙧᡂ㐣⛬䛻䛚䛡䜛 GroEL
䛾 Hinge䊡㒊䛻Ꮡᅾ䛩䜛 Gly192 ṧᇶ䛾ᙺ
GroEL
䛾Hinge䊡㒊䛿䜰䝢䜹䝹䝗䝯䜲䞁䛾ືస䛻㔜せ䛺ᙺ䜢ᯝ䛯䛧䠈䛣䛾㒊䛻Ꮡᅾ
䛧䛶䛔䜛Gly192
ṧᇶ䜢䛾䜰䝭䝜㓟ṧᇶ䛻⨨䛩䜛䛣䛸䛷䜰䝢䜹䝹䝗䝯䜲䞁䛾ື䛝䜢ㄪᩚ䛷䛝 䜛ྍ⬟ᛶ䛿๓⠇䛷♧䛧䛯䚹GroEL䛾䜰䝢䜹䝹䝗䝯䜲䞁䛷䛿䠈ᇶ㉁䛾⤖ྜ㒊䛸GroES
䛾⤖ྜ㒊 䛿㢮 ఝ 䛧䛯 ⨨ 䛻Ꮡ ᅾ 䛧䛶䛔䜛
(121)䚹Gly192
䜢 䛾䜰䝭䝜㓟 ṧ ᇶ 䛻⨨ 䛧䛯䛣䛸䛷Open
≧ែ䛻䛺䜛䛸GroEL G192X
ኚ␗య䛿ATP
䛾㠀Ꮡᅾୗ䛷GroES
䛜⤖ྜ䛩䜛䚹䛣䜜 䛻䜘䜚ኚᛶ≧ែ䛾MDH
䛸Rhodanese
䛿䝉䞁䝖䝷䝹䜻䝱䝡䝔䜱ෆ䛻ධ䜛䛣䛸䛜䛷䛝䛪䠈䝸䝣䜷䞊 䝹䝕䜱䞁䜾䜢ຓ䛷䛝䛺䛔䛣䛸䛜♧䛥䜜䛯䚹ANS⺯ග ᐃ䛾⤖ᯝ䛻䛚䛔䛶䠈Gly192ṧᇶ䜢⨨ 䛧䛯 䜰䝭䝜 㓟 ṧ ᇶ 䛜 䛝䛔䜋䛹
Open
≧ ែ 䛸 䛺䜚䜰䝢䜹䝹䝗䝯䜲䞁䛜 ⁐ ᾮ ୰ 䛻 ᭚ 䛥䜜䛯G192X
ኚ␗య䜋䛹⾲㠃Ỉᛶ䛜㧗䛟䛺䛳䛯䛣䛸䛛䜙䠈䛭䛾⾲㠃䛻Ỉᛶṧᇶ䜢ከ䛟ྵ䜣䛷䛔䜛ኳ↛ኚᛶ䝍䞁䝟䜽㉁䛸䜘䜚┦స⏝䛧䜔䛩䛔䛣䛸䛜⪃䛘䜙䜜䛯䚹⮅⮚䛛䜙ศἪ䛥䜜䜛䝨䝥䝏䝗 䝩䝹䝰䞁䛾୍䛴䛷䛒䜛䜲䞁䝇䝸䞁䛿䠈䜰䝭䝻䜲䝗⥺⥔◊✲䛾䝰䝕䝹䝍䞁䝟䜽㉁䛸䛧䛶䛥䜜䛶䛔䜛
(122)䚹䛣䜜䜎䛷䠈䜾䝹䞊䝥䊡ᆺ䝅䝱䝨䝻䝙䞁䛻ᒓ䛩䜛㉸㧗⇕ᛶ䛾ྂ⣽⳦⏤᮶䛾䝅䝱䝨䝻䝙䞁
(HSP60)䛾 C
ᮎ➃䛻䛚䛡䜛ᰂ㌾ᛶ䛜ቑຍ䛧䠈㧗 ᮲௳ୗ䛷䛺䛟䛸䜒㧗䛔άᛶ䜢♧䛩ኚ␗య䛻䛚䛔䛶䠈䜲䞁䝇䝸䞁䛜ᙧ ᡂ 䛩䜛⥺ ⥔ ᩿ ∦ 䛸⤖ ྜ 䛧䠈⥺ ⥔ 䜢ศ ゎ 䛩䜛䛣䛸䛜 ሗ ࿌ 䛥䜜䛶䛔䜛
(123)䚹䛣䛾 HSP60
䛿ATP
䛾Ꮡᅾୗ䛻䛚䛔䛶䜲䞁䝇䝸䞁⥺⥔䛾Thioflavin-T
⺯ග䛜ATP
㠀Ꮡᅾୗ䜘䜚䜒పୗ䛧䛶䛚䜚䠈ATPᏑᅾୗ䛷䃑䝅䞊䝖䛜䛥䜙䛻ῶᑡ䛧䛶䛔䜛䛾䛜♧䛥䜜䛶䛔䜛䚹 䛥䜙䛻䠈SHSY5Y ⣽⬊䜢⏝䛧䛯MTT
ศᯒ䛻䛚䛔䛶䠈 HSP60 䛸ATP
Ꮡᅾୗ䛻䛚䛔䛶䜰 䝭䝻䜲䝗⥺⥔䛿⬺จ㞟䛥䜜䠈⥺⥔䛾䜏䛸ẚ㍑䛧䛶≉䛻㧗䛔ẘᛶ⏘≀䜢ᙧᡂ䛧䛯䚹ATP 㠀Ꮡᅾୗ䛻䛚䛡䜛