Distinct insulin granule subpopulations implicated in the secretory pathology of diabetes types 1 and 2.

Insulin secretion from β-cells is reduced at the onset of type-1 and during type-2 diabetes. Although inflammation and metabolic dysfunction of β-cells elicit secretory defects associated with type-1 or type-2 diabetes, accompanying changes to insulin granules have not been established. To address this, we performed detailed functional analyses of insulin granules purified from cells subjected to model treatments that mimic type-1 and type-2 diabetic conditions and discovered striking shifts in calcium affinities and fusion characteristics.

Quiet Outer Membrane Protein G (OmpG) Nanopore for Biosensing.

Interest in nanopore technology has been growing due to nanopores' unique capabilities in small molecule sensing, measurement of protein folding, and low-cost DNA and RNA sequencing. The E. coli β-barrel outer membrane protein OmpG is an excellent alternative to other protein nanopores because of its single polypeptide chain.

A molecular mechanism for calcium-mediated synaptotagmin-triggered exocytosis.

The regulated exocytotic release of neurotransmitter and hormones is accomplished by a complex protein machinery whose core consists of SNARE proteins and the calcium sensor synaptotagmin-1. We propose a mechanism in which the lipid membrane is intimately involved in coupling calcium sensing to release. We found that fusion of dense core vesicles, derived from rat PC12 cells, was strongly linked to the angle between the cytoplasmic domain of the SNARE complex and the plane of the target membrane.

Quaternary structure of the small amino acid transporter OprG from .

is an opportunistic human pathogen that causes nosocomial infections. The outer membrane contains specific porins that enable substrate uptake, with the outer membrane protein OprG facilitating transport of small, uncharged amino acids. However, the pore size of an eight-stranded β-barrel monomer of OprG is too narrow to accommodate even the smallest transported amino acid, glycine, raising the question of how OprG facilitates amino acid uptake.

Reconstitution of calcium-mediated exocytosis of dense-core vesicles.

Regulated exocytosis is a process by which neurotransmitters, hormones, and secretory proteins are released from the cell in response to elevated levels of calcium. In cells, secretory vesicles are targeted to the plasma membrane, where they dock, undergo priming, and then fuse with the plasma membrane in response to calcium. The specific roles of essential proteins and how calcium regulates progression through these sequential steps are currently incompletely resolved.

OprG Harnesses the Dynamics of its Extracellular Loops to Transport Small Amino Acids across the Outer Membrane of Pseudomonas aeruginosa.

OprG is an outer membrane protein of Pseudomonas aeruginosa whose function as an antibiotic-sensitive porin has been controversial and not well defined. Circumstantial evidence led to the proposal that OprG might transport hydrophobic compounds by using a lateral gate in the barrel wall thought to be lined by three conserved prolines. To test this hypothesis and to find the physiological substrates of OprG, we reconstituted the purified protein into liposomes and found it to facilitate the transport of small amino acids such as glycine, alanine, valine, and serine, which was confirmed by Pseudomonas growth assays.

Highly parallel transport recordings on a membrane-on-nanopore chip at single molecule resolution.

Membrane proteins are prime drug targets as they control the transit of information, ions, and solutes across membranes. Here, we present a membrane-on-nanopore platform to analyze nonelectrogenic channels and transporters that are typically not accessible by electrophysiological methods in a multiplexed manner. The silicon chip contains 250,000 femtoliter cavities, closed by a silicon dioxide top layer with defined nanopores.

Layer-by-layer deposition of vesicles mediated by supramolecular interactions.

Vesicles are dynamic supramolecular structures with a bilayer membrane consisting of lipids or synthetic amphiphiles enclosing an aqueous compartment. Lipid vesicles have often been considered as mimics for biological cells. In this paper, we present a novel strategy for the preparation of three-dimensional multilayered structures in which vesicles containing amphiphilic β-cyclodextrin are interconnected by proteins using cyclodextrin guests as bifunctional linker molecules.

Tethered proteoliposomes containing human ABC transporter MRP3: new perspectives for biosensor application based on transmembrane proteins.

While transmembrane proteins and transporters comprise one of the largest protein families, their use in biosensors like biochips or lab-on-a-chip devices has so far been limited by their demanding requirements of a stable and compartmentalized lipid environment. A possible remedy lies in the tethering of proteoliposomes containing the reconstituted transmembrane protein to the biosensoric surface. As a proof of concept, we reconstituted the human ABC transporter MRP3 into biotinylated proteoliposomes and tethered those to a gold surface coated with streptavidin on a biotinylated self-assembled thiol monolayer.

Substrate translocation and stimulated ATP hydrolysis of human ABC transporter MRP3 show positive cooperativity and are half-coupled.

ABC transporters are involved in countless processes from lipid excretion over cellular detoxification to multidrug resistance of cancer cells. The latter is especially conferred by the ABCC subfamily also called multidrug resistance-associated proteins (MRPs) that excrete a variety of amphipathics including anticancer drugs by ATP-dependent transport. As the mechanisms of substrate translocation and ATP hydrolysis are still unclear for MRPs, we investigated the kinetics of both processes with focus on cooperativity and coupling between ATPase activity and substrate transport using purified MRP3 in proteoliposomes.

New synthetic procedures to catena-phosphorus cations: preparation and dissociation of the first cyclo-phosphino-halophosphonium salts.

Chlorination of 1,2,3,4-tetracyclohexyl-cyclo-tetraphosphine (2) by PhICl(2) or PCl(5) in the presence of Me(3)SiOTf or GaCl(3) provides a stepwise approach to salts of the first cyclo-phosphino-chlorophosphonium cations [Cy(4)P(4)Cl](+) ([19](+)) and [Cy(4)P(4)Cl(2)](2+) ([20](2+)). The analogous iodo derivative [Cy(4)P(4)I](+) ([17](+)) is obtained as the tetraiodogallate salt from reaction of 2 with I(2) in the presence of GaI(3). Reactions of the dication [20](2+) with PMe(3) or dmpe effect a dissociation of the cyclic framework resulting in the formation of salts containing [Me(3)PPCyPCyPMe(3)](2+) ([27](2+)), [dmpeCyP](2+) ([29](2+)), and [dmpeCyPCyP](2+) ([30](2+)), respectively.