LptB-LptF coupling mediates the closure of the substrate-binding cavity in the LptB2FGC transporter through a rigid-body mechanism to extract LPS

Lipopolysaccharides (LPS) are essential envelope components in many Gram-negative bacteria and provide intrinsic resistance to antibiotics. LPS molecules are synthesized in the inner membrane and then transported to the cell surface by the LPS transport (Lpt) machinery. In this system, the ATP-binding cassette (ABC) transporter LptB₂FGC extracts LPS from the inner membrane and places it onto a periplasmic protein bridge through a poorly understood mechanism. Here, we show that residue E86 of LptB is essential for coupling the function of this ATPase to that of its partners LptFG, specifically at the step where ATP binding drives the closure of the LptB dimer and the collapse of the LPS-binding cavity in LptFG that moves LPS to the Lpt periplasmic bridge. We also show that defects caused by changing residue E86 are suppressed by mutations altering either LPS structure or transmembrane helices in LptG. Furthermore, these suppressors also fix defects in the coupling helix of LptF, but not of LptG. Together, these results support a transport mechanism in which the ATP-driven movements of LptB and those of the substrate-binding cavity in LptFG are bi-directionally coordinated through the rigid-body coupling, with LptF’s coupling helix being important in coordinating cavity collapse with LptB dimerization.     

An Optimized Proteomics Approach Reveals Novel Alternative Proteins in Mouse Liver Development

Alternative ORFs (AltORFs) are unannotated sequences in genome that encode novel peptides or proteins named alternative proteins (AltProts). Although ribosome profiling and bioinformatics predict a large number of AltProts, mass spectrometry as the only direct way of identification is hampered by the short lengths and relative low abundance of AltProts. There is an urgent need for improvement of mass spectrometry methodologies for AltProt identification. Here, we report an approach based on size-exclusion chromatography for simultaneous enrichment and fractionation of AltProts from complex proteome. This method greatly simplifies the variance of AltProts discovery by enriching small proteins smaller than 40 kDa. In a systematic comparison between 10 methods, the approach we reported enabled the discovery of more AltProts with overall higher intensities, with less cost of time and effort compared to other workflows. We applied this approach to identify 89 novel AltProts from mouse liver, 39 of which were differentially expressed between embryonic and adult mice. During embryonic development, the upregulated AltProts were mainly involved in biological pathways on RNA splicing and processing, whereas the AltProts involved in metabolisms were more active in adult livers. Our study not only provides an effective approach for identifying AltProts but also novel AltProts that are potentially important in developmental biology.     

Enhancing native chemical ligation for challenging chemical protein syntheses

Native chemical ligation has enabled the chemical synthesis of proteins for a wide variety of applications (e.g., mirror-image proteins). However, inefficiencies of this chemoselective ligation in the context of large or otherwise challenging protein targets can limit the practical scope of chemical protein synthesis. In this review, we focus on recent developments aimed at enhancing and expanding native chemical ligation for challenging protein syntheses. Chemical auxiliaries, use of selenium chemistry, and templating all enable ligations at otherwise suboptimal junctions. The continuing development of these tools is making the chemical synthesis of large proteins increasingly accessible.     

“Our standpoint different from common...” (Scientific heritage of Alexander Gurwitsch)

The work of prominent Russian biologist Alexander Gavrilovich Gurwitsch (1874–1954) on the theory of organism development are reviewed. Alexander Gurwitsch introduced the concept of embryonic (morphogenetic, biological, and cellular) field and proposed several revisions of it from 1912 to 1944. Although neither of them can be considered as a final theory of development, his the persistent search for the invariant law that allows the shape (spatial structure) to be proposed for each next developmental stage from the previous shape is of imperishable methodological interest. Alexander Gurwitsch anticipated many ideas of the future theory of self-organization. His theoretical constructions are explicit and experiment-oriented but absolutely not esoteric. They represent a highly important and original contribution to theoretical biology and are an essential step to further development of the ontogenetic theory.     

Biogeography of the Limacoidea sensu lato (Gastropoda: Stylommatophora): vicariance events and long-distance dispersal

Aim Reconstruction of the historical biogeography of the Limacoidea sensu lato (including the Staffordiidae, Dyakiidae, Gastrodontoidea, Parmacelloidea, Zonitidae, Helicarionoidea and Limacoidea). Evaluation of the relative importance of dispersal and its consequences. Location World‐wide. Methods Weighted ancestral area analysis. Results The ancestral areas of the individual clades have been delimited using weighted ancestral area analysis and a sequence of possible vicariance and dispersal events has been suggested. The results of the ancestral area analysis have tentatively been correlated with Cretaceous and Tertiary palaeogeography. The widely overlapping distribution patterns of several families of the Limacoidea testify to extensive dispersal events. Dispersal capacity of land snails is correlated with body size. The significant negative correlation between body size and distribution area size corroborates the importance of passive dispersal for the evolution of the distribution patterns. Main conclusions The existence of extensive dispersal events of poor active dispersers like land snails diminishes the importance of recent distribution patterns for the reconstruction of palaeogeography. On the other hand, dispersal ensures that biogeographical data reflect the geographical configurations at a given time and renders the use of palaeobiogeographic data for the reconstruction of palaeogeographic configurations of the respective age possible.     

The seasonal dynamics of Copidodiaptomus numidicus (Gurney, 1909) and Thermocyclops dybowskii (Lande, 1890) in Castelo-do-Bode Reservoir

We investigated the population dynamics of Copidodiaptomus numidicus (Copepoda: Calanoida) and Thermocyclops dybowskii (Copepoda: Cyclopoida) in Castelo-do-Bode Reservoir (Portugal). Both species in the reservoir were regulated mainly by food availability during the summers of 1993 and 1994. C. numidicus was always more abundant than T. dybowskii in the reservoir. C. numidicus produced two generations during the sampling period of both years whereas T. dybowskii produced two and three generations in 1993 and 1994, respectively. Because of high temperatures and low rainfall in 1994, the reservoir was more eutrophic than in 1993. Higher clutch size and a higher percentage of ovigerous females suggest that both species were benefiting from better feeding conditions in 1994. Despite this, population growth was reduced in 1994 relatively to 1993. Vertebrate predation and predation by copepods seem to have been minor factors in explaining this decline. However, the presence of Mesostoma sp. in the reservoir may have contributed to the reduction of both copepod populations during specific periods in 1994 although the calanoid was more affected than the cyclopoid by this predation.