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Azide-alkyne cycloaddition reaction is a very common organic reaction to synthesize nitrogen-containing heterocycles. Once catalyzed by Cu(I) or Ru(II), it turns out to be a click reaction and thus is widely applied in chemical biology for labeling. However, besides their poor regioselectivity towards this reaction, these metal ions are not biologically friendly. Hence, it is an urgent need to develop a metal-free azide–alkyne cycloaddition reaction for biomedical applications. In this work, we found that, in the absence of metal ions, supramolecular self-assembly in an aqueous solution could realize this reaction with excellent regioselectivity. Nap-Phe-Phe-Lys(azido)-OH firstly self-assembled into nanofibers. Then, Nap-Phe-Phe-Gly(alkynyl)-OH at equivalent concentration approached to react with the assembly to yield the cycloaddition product Nap-Phe-Phe-Lys(triazole)-Gly-Phe-Phe-Nap to form nanoribbons. Due to space confinement effect, the product was obtained with excellent regioselectivity. Employing the excellent properties of supramolecular self-assembly, we are applying this strategy to realize more reactions without metal ion catalysis. Metal-free versions of azide–alkyne cycloadditions could find widespread applications in biomedical contexts. Here, the authors report an assembly-driven, regioselective azide–alkyne cycloaddition. [ABSTRACT FROM AUTHOR]

The S. cerevisiae plasma membrane H+-ATPase, Pma1, is a P3A-type ATPase and the primary protein component of the membrane compartment of Pma1 (MCP). Like other plasma membrane H+-ATPases, Pma1 assembles and functions as a hexamer, a property unique to this subfamily among the larger family of P-type ATPases. It has been unclear how Pma1 organizes the yeast membrane into MCP microdomains, or why it is that Pma1 needs to assemble into a hexamer to establish the membrane electrochemical proton gradient. Here we report a high-resolution cryo-EM study of native Pma1 hexamers embedded in endogenous lipids. Remarkably, we found that the Pma1 hexamer encircles a liquid-crystalline membrane domain composed of 57 ordered lipid molecules. The Pma1-encircled lipid patch structure likely serves as the building block of the MCP. At pH 7.4, the carboxyl-terminal regulatory α-helix binds to the phosphorylation domains of two neighboring Pma1 subunits, locking the hexamer in the autoinhibited state. The regulatory helix becomes disordered at lower pH, leading to activation of the Pma1 hexamer. The activation process is accompanied by a 6.7 Å downward shift and a 40° rotation of transmembrane helices 1 and 2 that line the proton translocation path. The conformational changes have enabled us to propose a detailed mechanism for ATP-hydrolysis-driven proton pumping across the plasma membrane. Our structures will facilitate the development of antifungal drugs that target this essential protein. The plasma membrane H+ -ATPase is responsible for maintenance of the plasma membrane potential, which provides energy for the transport of nutrients, and the plasma membrane H+ -ATPase in S. cerevisiae (Pma1) is a P3A-type ATPase that assembles and functions as a hexamer. Here, the authors present the cryo-EM structures of autoinhibited and activated native Pma1 hexamers purified with endogenous lipids and they propose a mechanism for proton pumping across the membrane by this family of H+ -ATPases. [ABSTRACT FROM AUTHOR]