Phosphine-alkene ligand-mediated alkyl-alkyl and alkyl-halide elimination processes from palladium(II)

N-Diphenylphosphino-7-aza-benzobicyclo[2.2.1]hept-2-ene (2) behaves as a chelating phosphine–alkene ligand for Pd0 and PdII, promoting direct alkyl–alkyl and indirect alkyl–halide reductive elimination reactions due to the stabilisation of the resulting bis(phosphine–alkene)Pd0 complex

Combined DFT and experimental studies of C–C and C–X elimination reactions promoted by a chelating phosphine–alkene ligand: the key role of penta-coordinate PdII

A combined computational and experimental study of the coordination chemistry of phosphine–alkene ligand L1 (N-diphenylphosphino-7-aza-benzobicyclo[2.2.1]hept-2-ene) with Pd0 and PdII is presented. Experimentally it is established that ligand L1 promotes direct alkyl–alkyl and indirect alkyl–halide reductive elimination from PdII species, affording the palladium(0) complex [Pd(κ2-P,C-L1)2] (2) in each case. The effectiveness of L1 in promoting these reactions is attributed to the initial formation of a penta-coordinate intermediate [PdMe(X)(κ1-P-L1)(κ2-P,C-L1)] (X = Me, Cl) coupled with the ease with which it transforms to 2. From computation, a lower activation barrier for C(sp3)–C(sp3) coupling and subsequent elimination has been computed for a stepwise associative pathway involving the initial formation of [PdMe2(κ1-P-L1)(κ2-P,C-L1)], compared to that computed for direct elimination from its parent, cis-[PdMe2(κ2-P,C-L1)]. Moreover, the C(sp3)–C(sp3) coupling reaction has been found to be primarily under thermodynamic control. It has also been demonstrated computationally that the methyl group of penta-coordinate [PdCl(Me)(κ1-P-L1)(κ2-P,C-L1)] is susceptible to nucleophilic attack by the phosphorus lone pair of a further equivalent of ligand L1, which proceeds through an SN2-like transition state. This initiates an unusual, indirect intermolecular reductive elimination process, resulting in the formation of equimolar quantities of the methyl phosphonium chloride salt of L1 and complex 2, in agreement with experimental observations. In contrast to the C(sp3)–C(sp3) coupling, computation shows that this indirect C(sp3)–Cl reductive elimination process is essentially under kinetic control

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field