The circular bacteriocin enterocin NKR-5-3B has an improved stability profile over nisin.

Bacteriocins are a large family of bacterial peptides that have antimicrobial activity and potential applications as clinical antibiotics or food preservatives. Circular bacteriocins are a unique class of these biomolecules distinguished by a seamless circular topology, and are widely assumed to be ultra-stable based on this constraining structural feature. However, without quantitative studies of their susceptibility to defined thermal, chemical, and enzymatic conditions, their stability characteristics remain poorly understood, limiting their translational development.

A Novel Pathway for Biosynthesis of the Herbicidal Phosphonate Natural Product Phosphonothrixin Is Widespread in Actinobacteria.

Phosphonothrixin is an herbicidal phosphonate natural product with an unusual, branched carbon skeleton. Bioinformatic analyses of the gene cluster, which is responsible for synthesis of the compound, suggest that early steps of the biosynthetic pathway, up to production of the intermediate 2,3-dihydroxypropylphosphonic acid (DHPPA) are identical to those of the unrelated phosphonate natural product valinophos. This conclusion was strongly supported by the observation of biosynthetic intermediates from the shared pathway in spent media from two phosphonothrixin producing strains.

BID-seq: The Quantitative and Base-Resolution Sequencing Method for RNA Pseudouridine.

Quantitative and base-resolution sequencing methods are critical to investigations of the biological functions of diverse RNA modifications. These methods may also be employed for clinical studies and clinical applications in the future. In this In Focus article, we introduce and discuss the development of Bisulfite-Induced Deletion sequencing (BID-seq) for quantitatively detecting mRNA pseudouridine (Ψ) modifications at base resolution.

Genome-wide Analysis Reflects Novel 5-Hydroxymethylcytosines Implicated in Diabetic Nephropathy and the Biomarker Potential.

Aim: Diabetic nephropathy (DN) has become the most common cause of end-stage renal disease (ESRD) in most countries. Elucidating novel epigenetic contributors to DN can not only enhance our understanding of this complex disorder, but also lay the foundation for developing more effective monitoring tools and preventive interventions in the future, thus contributing to our ultimate goal of improving patient care.

Methods: The 5hmC-Seal, a highly selective, chemical labeling technique, was used to profile genome-wide 5-hydroxymethylcytosines (5hmC), a stable cytosine modification type marking gene activation, in circulating cell-free DNA (cfDNA) samples from a cohort of patients recruited at Zhongnan Hospital, including T2D patients with nephropathy (DN, n = 12), T2D patients with non-DN vascular complications (non-DN, n = 29), and T2D patients without any complication (controls, n = 14).

Macrocyclization and Backbone Modification in RiPP Biosynthesis.

The past decade has seen impressive advances in understanding the biosynthesis of ribosomally synthesized and posttranslationally modified peptides (RiPPs). One of the most common modifications found in these natural products is macrocyclization, a strategy also used by medicinal chemists to improve metabolic stability and target affinity and specificity. Another tool of the peptide chemist, modification of the amides in a peptide backbone, has also been observed in RiPPs.

Engineering of new-to-nature ribosomally synthesized and post-translationally modified peptide natural products.

Natural products have historically been important lead sources for drug development, particularly to combat infectious diseases. Increasingly, their structurally complex scaffolds are also envisioned as leads for applications for which they did not evolve, an approach aided by engineering of new-to-nature analogs. Ribosomally synthesized and post-translationally modified peptides (RiPPs) are promising candidates for bioengineering because they are genetically encoded and their biosynthetic enzymes display significant substrate tolerance.

Cameo appearances of aminoacyl-tRNA in natural product biosynthesis.

The breadth of unprecedented enzymatic reactions performed during the formation of microbial natural products has continued to expand as new biosynthetic gene clusters are unearthed by genome mining. Enzymes that use aminoacyl-tRNA (aa-tRNA) outside of the translation machinery have been known for decades, and accounts of their use in natural product biosynthesis are just beginning to accumulate. This review will highlight the recent discoveries and advances in our mechanistic understanding of aa-tRNA-dependent enzymes that play key roles in the biosynthesis of a growing number of microbial natural products.

An oxidative fluctuation hypothesis of aging generated by imaging H₂O₂ levels in live Caenorhabditis elegans with altered lifespans.

Reactive oxygen species (ROS) are important factors mediating aging according to the free radical theory of aging. Few studies have systematically measured ROS levels in relationship to aging, partly due to the lack of tools for detection of specific ROS in live animals. By using the H₂O₂-specific fluorescence probe Peroxy Orange 1, we assayed the H₂O₂ levels of live Caenorhabditis elegans with 41 aging-related genes being individually knocked down by RNAi.

Discovery and biosynthesis of phosphonate and phosphinate natural products.

The P-C bonds in phosphonate and phosphinate natural products endow them with a high level of stability and the ability to mimic phosphate esters and carboxylates. As such, they have a diverse range of enzyme targets that act on substrates containing such functionalities. Recent years have seen a renewed interest in discovery efforts focused on this class of compounds as well as in understanding their biosynthetic pathways.

A selective reaction-based fluorescent probe for detecting cobalt in living cells.

A reaction-based strategy exploiting cobalt-mediated oxidative C-O bond cleavage affords a selective turn-on fluorescent probe for paramagnetic Co(2+) in water and in living cells.

A two-photon fluorescent probe for ratiometric imaging of hydrogen peroxide in live tissue.

We report a two-photon fluorescent probe (PN1) that can be excited by 750 nm femto-second pulses, shows high photostability and negligible toxicity, and can visualize H(2)O(2) distribution in live cells and tissue by two-photon microscopy.

Lanthanide-based luminescent probes for selective time-gated detection of hydrogen peroxide in water and in living cells.

Lanthanide-based luminescent probes TPR1 and TPR2 were developed for the detection of hydrogen peroxide (H(2)O(2)) in living systems. The chemoselective reaction of these boronate-protected probes with H(2)O(2) resulted in an enhanced lanthanide sensitization and a 6-fold increase in luminescent intensity. TPR2 was utilized to measure the endogenous production of H(2)O(2) in RAW 264.

Visualizing ascorbate-triggered release of labile copper within living cells using a ratiometric fluorescent sensor.

We present the synthesis, properties, and biological applications of Ratio-Coppersensor-1 (RCS1), a new water-soluble fluorescent sensor for ratiometric imaging of copper in living cells. RCS1 combines an asymmetric BODIPY reporter and thioether-based ligand receptor to provide high selectivity and sensitivity for Cu(+) over other biologically relevant metal ions, including Cu(2+) and Zn(2+), a ca. 20-fold fluorescence ratio change upon Cu(+) binding, and visible excitation and emission profiles compatible with standard fluorescence microscopy filter sets.

A turn-on fluorescent sensor for detecting nickel in living cells.

We present the synthesis and properties of Nickelsensor-1 (NS1), a new water-soluble, turn-on fluorescent sensor that is capable of selectively responding to Ni(2+) in aqueous solution and in living cells. NS1 combines a BODIPY chromophore and a mixed N/O/S receptor to provide good selectivity for Ni(2+) over a range of biologically abundant metal ions in aqueous solution. In addition to these characteristics, confocal microscopy experiments further show that NS1 can be delivered into living cells and report changes in intracellular Ni(2+) levels in a respiratory cell model.

Chapter 21. In vitro studies of lantibiotic biosynthesis.

The lantibiotics are ribosomally synthesized and posttranslationally modified peptide antibiotics containing the thioether crosslinks lanthionine (Lan) and 3-methyllanthionine (MeLan) and typically also the dehydroamino acids dehydroalanine (Dha) and (Z)-dehydrobutyrine (Dhb). These modifications are formed by dehydration of Ser/Thr residues to produce the Dha and Dhb structures, and subsequent conjugate additions of Cys residues onto the unsaturated amino acids to form thioether rings (Lan and MeLan). Several of the enzymatic reactions involved in lantibiotic biosynthesis have been reconstituted in vitro in recent years and these systems as well as a general overview of lantibiotic biosynthesis are discussed in this chapter.