The contribution of achiral residues in the laspartomycin family of calcium-dependent lipopeptide antibiotics

The growing threat of antibacterial resistance is a global concern. The so-called calcium-dependent lipopeptide antibiotics (CDAs) have emerged as a promising source of new antibiotic agents that are rich in structural and mechanistic diversity. Over forty unique CDAs have been identified to date and share a number of common features. Recent efforts in our group have provided new mechanistic and structural insights into the laspartomycin family of CDAs. We here describe investigations aimed at probing the role of the three glycine residues found in the laspartomycin peptide macrocycle. In doing so laspartomycin analogues containing the achiral 2-aminoisobutyric acid (AIB) as well as L- or D-alanine in place of glycine were prepared and their antibacterial activities evaluated.Microbial Biotechnolog

A convenient chemoenzymatic preparation of chimeric macrocyclic peptide antibiotics with potent activity against gram-negative pathogens

Microbial Biotechnolog

Performance Enhancement of Electrocatalytic Hydrogen Evolution through Coalescence-Induced Bubble Dynamics

The evolution of electrogenerated gas bubbles during water electrolysis can significantly hamper the overall process efficiency. Promoting the departure of electrochemically generated bubbles during (water) electrolysis is therefore beneficial. For a single bubble, a departure from the electrode surface occurs when buoyancy wins over the downward-acting forces (e.g., contact, Marangoni, and electric forces). In this work, the dynamics of a pair of H2 bubbles produced during the hydrogen evolution reaction in 0.5 M H2SO4 using a dual platinum microelectrode system is systematically studied by varying the electrode distance and the cathodic potential. By combining high-speed imaging and electrochemical analysis, we demonstrate the importance of bubble-bubble interactions in the departure process. We show that bubble coalescence may lead to substantially earlier bubble departure as compared to buoyancy effects alone, resulting in considerably higher reaction rates at a constant potential. However, due to continued mass input and conservation of momentum, repeated coalescence events with bubbles close to the electrode may drive departed bubbles back to the surface beyond a critical current, which increases with the electrode spacing. The latter leads to the resumption of bubble growth near the electrode surface, followed by buoyancy-driven departure. While less favorable at small electrode spacing, this configuration proves to be very beneficial at larger separations, increasing the mean current up to 2.4 times compared to a single electrode under the conditions explored in this study.</p

The origin and rise of complex life:progress requires interdisciplinary integration and hypothesis testing

Understanding of the triggers and timing of the rise of complex life ca 2100 to 720 million years ago has expanded dramatically in recent years. This theme issue brings together diverse and novel geochemical and palaeontological data presented as part of the Royal Society ‘The origin and rise of complex life: integrating models, geochemical and palaeontological data’ discussion meeting held in September 2019. The individual papers offer prescient insights from multiple disciplines. Here we summarize their contribution towards the goal of the meeting; to create testable hypotheses for the differing roles of changing climate, oceanic redox, nutrient availability, and ecosystem feedbacks across this profound, but enigmatic, transitional period

A β-hairpin epitope as novel structural requirement for protein arginine rhamnosylation

For canonical asparagine glycosylation, the primary amino acid sequence that directs glycosylation at specific asparagine residues is well-established. Here we reveal that a recently discovered bacterial enzyme EarP, that transfers rhamnose to a specific arginine residue in its acceptor protein EF-P, specifically recognizes a beta-hairpin loop. Notably, while the in vitro rhamnosyltransferase activity of EarP is abolished when presented with linear substrate peptide sequences derived from EF-P, the enzyme readily glycosylates the same sequence in a cyclized beta-hairpin mimic. Additional studies with other substrate-mimicking cyclic peptides revealed that EarP activity is sensitive to the method used to induce cyclization and in some cases is tolerant to amino acid sequence variation. Using detailed NMR approaches, we established that the active peptide substrates all share some degree of beta-hairpin formation, and therefore conclude that the beta-hairpin epitope is the major determinant of arginine-rhamnosylation by EarP. Our findings add a novel recognition motif to the existing knowledge on substrate specificity of protein glycosylation, and are expected to guide future identifications of rhamnosylation sites in other protein substrates

The cell envelope stress response of bacillus subtilis towards laspartomycin C

Microbial Biotechnolog

Binding studies reveal phospholipid specificity and its role in the calcium-dependent mechanism of action of daptomycin

Multidrug-resistant bacteria pose a serious global health threat as antibiotics are increasingly losing their clinical efficacy. A molecular level understanding of the mechanism of action of antimicrobials plays a key role in developing new agents to combat the threat of antimicrobial resistance. Daptomycin, the only clinically used calcium-dependent lipopeptide antibiotic, selectively disrupts Gram-positive bacterial membranes to illicit its bactericidal effect. In this study, we use isothermal titration calorimetry to further characterize the structural features of the target bacterial phospholipids that drive daptomycin binding. Our studies reveal that daptomycin shows a clear preference for the phosphoglycerol headgroup. Furthermore, unlike other calcium-dependent lipopeptide antibiotics, calcium binding by daptomycin is strongly dependent on the presence of phosphatidylglycerol. These investigations provide new insights into daptomycin's phospholipid specificity and calcium binding behavior.Microbial Biotechnolog

Thrombin-derived peptides potentiate the activity of Gram-positive-specific antibiotics against Gram-negative bacteria

Microbial Biotechnolog

Mechanistic insights into the C₅₅-P targeting lipopeptide antibiotics revealed by structure-activity studies and high-resolution crystal structures

AbstractThe continued rise of antibiotic resistance is a global concern that threatens to undermine many aspects of modern medical practice. Key to addressing this threat is the discovery and development of new antibiotics that operate by unexploited modes of action. The so-called calcium-dependent lipopeptide antibiotics (CDAs) are an important emerging class of natural products that provides a source of new antibiotic agents rich in structural and mechanistic diversity. Notable in this regard is the subset of CDAs comprising the laspartomycins and amphomycins/friulimicins that specifically target the bacterial cell wall precursor undecaprenyl phosphate (C55-P). In this study we describe the design and synthesis of new C55-P-targeting CDAs with structural features drawn from both the laspartomycin and amphomycin/friulimicin classes. Assessment of these lipopeptides revealed previously unknown and surprisingly subtle structural features that are required for antibacterial activity. High-resolution crystal structures further indicate that the amphomycin/friulimicin-like lipopeptides adopt a unique crystal packing that governs their interaction with C55-P and provides an explanation for their antibacterial effect. In addition, live-cell microscopy studies provide further insights into the biological activity of the C55-P targeting CDAs highlighting their unique mechanism of action relative to the clinically used CDA daptomycin.Microbial Biotechnolog