Binding of subunit E into the A–B interface of the A₁A_O ATP synthase

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• 作者：Cornelia Hunke^a ; Martin Antosch^b ; ¹ ; Volker M&#xfc ; ller^b ; Gerhard Gr&#xfc ; ber^a ; ^{ggrueber@ntu.edu.sg"" rel=""nofollow}
• 关键词：Archaeal ATP synthase ; A₁A_O ATP synthase ; Methanosarcina mazei Gö ; 1 ; Peripheral stalk ; Fluorescence correlation spectroscopy ; F₁F_O ATP synthase
• 刊名：Biochimica et Biophysica Acta (BBA)/Biomembranes
• 出版年：2011
• 出版时间：September 2011
• 年：2011
• 卷：1808
• 期：9
• 页码：2111-2118
• 全文大小：1019 K

文摘

Two of the distinct diversities of the engines A₁A_O ATP synthase and F₁F_O ATP synthase are the existence of two peripheral stalks and the 24 kDa stalk subunit E inside the A₁A_O ATP synthase. Crystallographic structures of subunit E have been determined recently, but the epitope(s) and the strength to which this subunit does bind in the enzyme complex are still a puzzle. Using the recombinant A₃B₃D complex and the major subunits A and B of the methanogenic A₁A_O ATP synthase in combination with fluorescence correlation spectroscopy (FCS) we demonstrate, that the stalk subunit E does bind to the catalytic headpiece formed by the A₃B₃ hexamer with an affinity (K_d) of 6.1 ± 0.2 μM. FCS experiments with single A and B, respectively, demonstrated unequivocally that subunit E binds stronger to subunit B (K_d = 18.9 ± 3.7 μM) than to the catalytic A subunit (K_d = 53.1 ± 4.4). Based on the crystallographic structures of the three subunits A, B and E available, the arrangement of the peripheral stalk subunit E in the A–B interface has been modeled, shining light into the A–B–E assembly of this enzyme.