Biocatalytic cyclization of small macrolactams by a penicillin-binding protein-type thioesterase.

Macrocyclic peptides represent promising scaffolds for chemical tools and potential therapeutics. Synthetic methods for peptide macrocyclization are often hampered by C-terminal epimerization and oligomerization, leading to difficult scalability. While chemical strategies to circumvent this issue exist, they often require specific amino acids to be present in the peptide sequence. Herein, we report the characterization of Ulm16, a peptide cyclase belonging to the penicillin-binding protein-type class of thioesterases that catalyze head-to-tail macrolactamization of nonribosmal peptides. Ulm16 efficiently cyclizes various nonnative peptides ranging from 4 to 6 amino acids with catalytic efficiencies of up to 3 × 106 M-1 s-1. Unlike many previously described homologs, Ulm16 tolerates a variety of C- and N-terminal amino acids. The crystal structure of Ulm16, along with modeling of its substrates and site-directed mutagenesis, allows for rationalization of this wide substrate scope. Overall, Ulm16 represents a promising tool for the biocatalytic production of macrocyclic peptides.

Electroanalysis and Spectroelectrochemistry of Nonaromatic Explosives in Acetonitrile Containing Dissolved Oxygen.

In search of a rapid, low-cost, and solution-phase detection technique for explosives, the (spectro-)electrochemistry of compounds from two major nonaromatic classes, namely nitramines (RDX and HMX) and nitrate esters (pentaerythritol tetranitrate (PETN) and the plastic explosive composite Semtex 1A) in acetonitrile (AN) is reported. In electrochemical screening, 5 µg of explosive material was detectable in 10 s by multicomponent cyclic voltammetric (CV) analysis on unmodified glassy carbon under ubiquitous environmental influences (i.e., trace water and dissolved oxygen). The explosives were identified with high recoveries under a battery of proof-of-concept testing scenarios in various matrices. In AN containing naturally dissolved oxygen (approx. 2 mM), the superoxide radical is co-electrogenerated during analyte reduction. Free superoxide yields prominent signals that the explosives attenuate quantitatively. To gain further insight into the electrochemical transformation mechanism, spectroelectrochemistry was employed to monitor changes in ultraviolet (UV) absorbance during CV and identify transient intermediates and product species, which could be targeted by future chemical sensors. Overlapping UV spectra of multiple species are deconvoluted using a new strategy, spectral regional baselining, for time- and potential-resolved spectroelectrochemical (SEC) analysis. This study shows that dissolved oxygen, hitherto an interferent purposefully removed from the solution, can be exploited advantageously in electrochemical sensing. The work expands our understanding of high-explosive solution-phase chemistry and offers a novel route to signal transduction for the sensing of energetic materials.

Synthetic Natural Product Inspired Cyclic Peptides.

Natural products are a bountiful source of bioactive molecules. Unfortunately, discovery of novel bioactive natural products is challenging due to cryptic biosynthetic gene clusters, low titers, and arduous purifications. Herein, we describe SNaPP (Synthetic Natural Product Inspired Cyclic Peptides), a method for identifying NP-inspired bioactive peptides. SNaPP expedites bioactive molecule discovery by combining bioinformatics predictions of nonribosomal peptide synthetases with chemical synthesis of the predicted natural products (pNPs). SNaPP utilizes a recently discovered cyclase, the penicillin binding protein-like cyclase, as the lynchpin for the development of a library of head-to-tail cyclic peptide pNPs. Analysis of 500 biosynthetic gene clusters allowed for identification of 131 novel pNPs. Fifty-one diverse pNPs were synthesized using solid phase peptide synthesis and solution-phase cyclization. Antibacterial testing revealed 14 pNPs with antibiotic activity, including activity against multidrug-resistant Gram-negative bacteria. Overall, SNaPP demonstrates the power of combining bioinformatics predictions with chemical synthesis to accelerate the discovery of bioactive molecules.