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Abstract Hyperactive Ras signalling is found in most cancers. Ras proteins are only active in membrane nanoclusters, which are therefore potential drug targets. We previously showed that the nanocluster scaffold galectin-1 (Gal1) enhances H-Ras nanoclustering via direct interaction with the Ras binding domain (RBD) of Raf. Here, we establish that the B-Raf preference of Gal1 emerges from the divergence of the Raf RBDs at their proposed Gal1-binding interface. We then identify the L5UR peptide, which disrupts this interaction by binding with low micromolar affinity to the B- and C-Raf-RBDs. Its 23-mer core fragment is sufficient to interfere with H-Ras nanoclustering, modulate Ras-signalling and moderately reduce cell viability. These latter two phenotypic effects may also emerge from the ability of L5UR to broadly engage with several RBD- and RA-domain containing Ras interactors. The L5UR-peptide core fragment is a starting point for the development of more specific reagents against Ras-nanoclustering and -interactors.

Targeting the interaction between transcription factor TEAD and its co-repressor VGL4 is an attractive strategy to chemically modulate Hippo signaling. Here, the authors develop a proteomimetic with stabilized tertiary structure that inhibits the TEAD:VGL4 interaction in vitro and in cells.

The rising incidence of multidrug resistance in Gram-negative bacteria underlines the urgency for novel treatment options. One promising new approach is the synergistic combination of antibiotics with antimicrobial peptides. However, the use of such peptides is not straightforward; they are often sensitive to proteolytic degradation, which greatly limits their clinical potential. One approach to increase stability is to apply a hydrocarbon staple to the antimicrobial peptide, thereby fixing them in an α-helical conformation, which renders them less exposed to proteolytic activity. In this work we applied several different hydrocarbon staples to two previously described peptides shown to act on the outer membrane, L6 and L8, and tested their activity in a zebrafish embryo infection model using a clinical isolate of Acinetobacter baumannii as a pathogen. We show that the introduction of such a hydrocarbon staple to the peptide L8 improves its in vivo potentiating activity on antibiotic treatment, without increasing its in vivo antimicrobial activity, toxicity or hemolytic activity.

The N-end rule pathway targets substrate proteins for proteasomal degradation. Here, Whiteet al. show that ArabidopsisPLANT CYSTEINE OXIDASEs show dioxygenase activity producing Cys-sulfinic acid at the N-terminus of target proteins, which then act as direct substrates for arginyl transferase.

Bicyclic peptides can inhibit biological targets hard to address with small molecules. Here, the authors combine two orthogonal ring-closing reactions to produce bicyclic peptides with improved bioactivity thereby providing a strategy that can greatly improve the structural diversity of such peptides.

ABSTRACT Most bacteria and archaea use the tubulin homologue FtsZ as its central organizer of cell division. In Gram-negative Escherichia coli bacteria, FtsZ recruits cytosolic, transmembrane, periplasmic, and outer membrane proteins, assembling the divisome that facilitates bacterial cell division. One such divisome component, FtsQ, a bitopic membrane protein with a globular domain in the periplasm, has been shown to interact with many other divisome proteins. Despite its otherwise unknown function, it has been shown to be a major divisome interaction hub. Here, we investigated the interactions of FtsQ with FtsB and FtsL, two small bitopic membrane proteins that act immediately downstream of FtsQ. We show in biochemical assays that the periplasmic domains of E. coli FtsB and FtsL interact with FtsQ, but not with each other. Our crystal structure of FtsB bound to the β domain of FtsQ shows that only residues 64 to 87 of FtsB interact with FtsQ. A synthetic peptide comprising those 24 FtsB residues recapitulates the FtsQ-FtsB interactions. Protein deletions and structure-guided mutant analyses validate the structure. Furthermore, the same structure-guided mutants show cell division defects in vivo that are consistent with our structure of the FtsQ-FtsB complex that shows their interactions as they occur during cell division. Our work provides intricate details of the interactions within the divisome and also provides a tantalizing view of a highly conserved protein interaction in the periplasm of bacteria that is an excellent target for cell division inhibitor searches. IMPORTANCE In most bacteria and archaea, filaments of FtsZ protein organize cell division. FtsZ forms a ring structure at the division site and starts the recruitment of 10 to 20 downstream proteins that together form a multiprotein complex termed the divisome. The divisome is thought to facilitate many of the steps required to make two cells out of one. FtsQ and FtsB are part of the divisome, with FtsQ being a central hub, interacting with most of the other divisome components. Here we show for the first time in detail how FtsQ interacts with its downstream partner FtsB and show that mutations that disturb the interface between the two proteins effectively inhibit cell division.