Atomically precise single-crystal structures of electrically conducting 2D metal-organic frameworks.

Electrically conducting 2D metal-organic frameworks (MOFs) have attracted considerable interest, as their hexagonal 2D lattices mimic graphite and other 2D van der Waals stacked materials. However, understanding their intrinsic properties remains a challenge because their crystals are too small or of too poor quality for crystal structure determination. Here, we report atomically precise structures of a family of 2D π-conjugated MOFs derived from large single crystals of sizes up to 200 μm, allowing atomic-resolution analysis by a battery of high-resolution diffraction techniques.

Quantifying the heterogeneity of macromolecular machines by mass photometry.

Sample purity is central to in vitro studies of protein function and regulation, and to the efficiency and success of structural studies using techniques such as x-ray crystallography and cryo-electron microscopy (cryo-EM). Here, we show that mass photometry (MP) can accurately characterize the heterogeneity of a sample using minimal material with high resolution within a matter of minutes. To benchmark our approach, we use negative stain electron microscopy (nsEM), a popular method for EM sample screening.

Atomic structure and enzymatic insights into the vancomycin-resistant Enterococcus faecalis (V583) alkylhydroperoxide reductase subunit C.

The Enterococcus faecalis alkyl hydroperoxide reductase complex (AhpR) with its subunits AhpC (EfAhpC) and AhpF (EfAhpF) are of paramount importance to restore redox homeostasis. Recently, the novel phenomenon of swapping of the catalytic domains of EfAhpF was uncovered. Here, we visualized its counterpart EfAhpC (187 residues) from the vancomycin-resistant E.

Structure, mechanism and ensemble formation of the alkylhydroperoxide reductase subunits AhpC and AhpF from Escherichia coli.

Hydroperoxides are reactive oxygen species (ROS) that are toxic to all cells and must be converted into the corresponding alcohols to alleviate oxidative stress. In Escherichia coli, the enzyme primarily responsible for this reaction is alkylhydroperoxide reductase (AhpR). Here, the crystal structures of both of the subunits of EcAhpR, EcAhpF (57 kDa) and EcAhpC (21 kDa), have been solved.

Key roles of the Escherichia coli AhpC C-terminus in assembly and catalysis of alkylhydroperoxide reductase, an enzyme essential for the alleviation of oxidative stress.

2-Cys peroxiredoxins (Prxs) are a large family of peroxidases, responsible for antioxidant function and regulation in cell signaling, apoptosis and differentiation. The Escherichia coli alkylhydroperoxide reductase (AhpR) is a prototype of the Prxs-family, and is composed of an NADH-dependent AhpF reductase (57 kDa) and AhpC (21 kDa), catalyzing the reduction of H2O2. We show that the E.

The N termini of a-subunit isoforms are involved in signaling between vacuolar H+-ATPase (V-ATPase) and cytohesin-2.

Previously, we reported an acidification-dependent interaction of the endosomal vacuolar H(+)-ATPase (V-ATPase) with cytohesin-2, a GDP/GTP exchange factor (GEF), suggesting that it functions as a pH-sensing receptor. Here, we have studied the molecular mechanism of signaling between the V-ATPase, cytohesin-2, and Arf GTP-binding proteins. We found that part of the N-terminal cytosolic tail of the V-ATPase a2-subunit (a2N), corresponding to its first 17 amino acids (a2N(1-17)), potently modulates the enzymatic GDP/GTP exchange activity of cytohesin-2.

Solution structure of subunit a, a₁₀₄₋₃₆₃, of the Saccharomyces cerevisiae V-ATPase and the importance of its C-terminus in structure formation.

The 95 kDa subunit a of eukaryotic V-ATPases consists of a C-terminal, ion-translocating part and an N-terminal cytosolic domain. The latter's N-terminal domain (~40 kDa) is described to bind in an acidification-dependent manner with cytohesin-2 (ARNO), giving the V-ATPase the putative function as pH-sensing receptor. Recently, the solution structure of the very N-terminal segment of the cytosolic N-terminal domain has been solved.

N-terminal domain of the V-ATPase a2-subunit displays integral membrane protein properties.

V-ATPase is a multisubunit membrane complex that functions as nanomotor coupling ATP hydrolysis with proton translocation across biological membranes. Recently, we uncovered details of the mechanism of interaction between the N-terminal tail of the V-ATPase a2-subunit isoform (a2N(1-402)) and ARNO, a GTP/GDP exchange factor for Arf-family small GTPases. Here, we describe the development of two methods for preparation of the a2N(1-402) recombinant protein in milligram quantities sufficient for further biochemical, biophysical, and structural studies.

The genetic interactome of prohibitins: coordinated control of cardiolipin and phosphatidylethanolamine by conserved regulators in mitochondria.

Prohibitin ring complexes in the mitochondrial inner membrane regulate cell proliferation as well as the dynamics and function of mitochondria. Although prohibitins are essential in higher eukaryotes, prohibitin-deficient yeast cells are viable and exhibit a reduced replicative life span. Here, we define the genetic interactome of prohibitins in yeast using synthetic genetic arrays, and identify 35 genetic interactors of prohibitins (GEP genes) required for cell survival in the absence of prohibitins.