Antimicrobial activity of simplified mimics of celogentin C

Linear and bicyclic analogues of the peptide natural product, celogentin C, have been prepared in which the sidechain–sidechain crosslinks in celogentin are omitted or replaced with a mesitylenyl moiety. The simplified bicyclic peptides display moderate antibacterial activity, potentially through inhibition of bacterial protomicrotubule formation, while the linear analogs show higher antibacterial activity through a possible membrane disruption mechanism.     

A comparison of positive and negative ion collision‐induced dissociation for model heptapeptides with one basic residue

The effects of the identity and position of basic residues on peptide dissociation were explored in the positive and negative modes. Low‐energy collision‐induced dissociation (CID) was performed on singly protonated and deprotonated heptapeptides of the type: XAAAAAA, AAAXAAA, AAAAAXA and AAAAAAX, where X is arginine (R), lysine (K) or histidine (H) residues and A is alanine. For [M + H]⁺, the CID spectra are dominated by cleavages adjacent to the basic residues and the majority of the product ions contain the basic residues. The order of a basic residue's influence on fragmentation of [M + H]⁺ is arginine > histidine ≈ lysine, which is also the order of decreasing gas‐phase basicity for these amino acids. These results are consistent with the side chains of basic residues being positive ion charge sites and with the more basic arginine residues having a higher retention (i.e. sequestering) of the positive charge. In contrast, for [M ? H]^? the identity and position of basic residues has almost no effect on backbone fragmentation. This is consistent with basic residues not being negative mode charge sites. For these peptides, more complete series of backbone fragments, which are important in the sequencing of unknowns, can be found in the negative mode. Spectra at both polarities contain C‐terminal y‐ions, but y_n″⁺ has two more hydrogens than the corresponding y_n^?. Another major difference is the production of the N‐terminal backbone series b_n⁺ in the positive mode and c_n^? in the negative mode. Thus, comparison of positive and negative ion spectra with an emphasis on searching for pairs of ions that differ by 2 Da (y_n″⁺ vs y_n^?) and by 15 Da (b_n⁺ vs c_n^?) may be a useful method for determining whether a product ion is generated from the C‐terminal or the N‐terminal end of a peptide. In addition, a characteristic elimination of NH?C?NH from arginine residues is observed for deprotonated peptides. Copyright © 2010 John Wiley & Sons, Ltd.     

Solid-phase synthesis of the cyclic lipononadepsipeptide [N-Mst(Ser1), d-Ser4, L-Thr6, L-Asp8, L-Thr9]syringotoxin

An optimized solid-phase strategy for the preparation of the cyclic lipononadepsipeptide [N-Mst(L-Ser1), D-Ser4, L-Thr6, L-Asp8, L-Thr9]syringotoxin is reported. The strategy is based on the use of a mild orthogonal protection scheme and the incorporation of the nonproteinogenic amino acid (Z)-Dhb into the peptide chain as the dipeptide Fmoc-Thr(tBu)-(Z)-Dhb-OH. The didehydrodipeptide was synthesized by a water-soluble carbodiimide-induced beta-elimination of a protected dipeptide containing a residue of Thr with its free hydroxy side chain unprotected.     

A straight forward and first total synthesis of Penilumamides B–D

A simple and straight forward first total synthesis of rare lumazine peptides, Penilumamide B, C and D isolated from the marine-derived fungi Aspergillus and Penicillium sp. is described from a common starting material, 1,3-dimethyllumazine-6-carboxylic acid. Penilumamide C was prepared from Penilumamide B by oxidation of the methionine residue to the corresponding sulphone.     

Evaluating the Properties of Ginger Protease-Degraded Collagen Hydrolysate and Identifying the Cleavage Site of Ginger Protease by Using an Integrated Strategy and LC-MS Technology

(1) Methods: An integrated strategy, including in vitro study (degree of hydrolysis (DH) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity) and in vivo study (absorption after oral administration in rats), was developed to evaluate the properties of the fish skin gelatin hydrolysates prepared using different proteases (pepsin, alkaline protease, bromelain, and ginger protease). Meanwhile, in order to identify the hydrolysis site of ginger protease, the peptides in the ginger protease-degraded collagen hydrolysate (GDCH) were comprehensively characterized by liquid chromatography/tandem mass spectrometry (LC-MS) method. (2) Results: The GDCH exhibited the highest DH (20.37%) and DPPH radical scavenging activity (77.73%), and in vivo experiments showed that the GDCH was more efficiently absorbed by the gastrointestinal tract. Further oral administration experiments revealed that GDCH was not entirely degraded to free amino acids and can be partially absorbed as dipeptides and tripeptides in intact forms, including Pro-Hyp, Gly-Pro-Hyp, and X-Hyp-Gly tripeptides. LC-MS results determined the unique substrate specificity of ginger protease recognizing Pro and Hyp at the P₂ position based on the amino acids at the P₂ position from the three types of tripeptides (Gly-Pro-Y, X-Hyp-Gly, and Z-Pro-Gly) and 136 identified peptides (>4 amino acids). Interestingly, it suggested that ginger protease can also recognize Ala in the P₂ position. (3) Conclusions: This study comprehensively evaluated the properties of GDCH by combining in vitro and in vivo strategies, and is the first to identify the cleavage site of ginger protease by LC-MS technique. It provides support for the follow-up study on the commercial applications of ginger protease and bioactivities of the hydrolysate produced by ginger protease.     

Peptaibol-Containing Extracts of Trichoderma atroviride and the Fight against Resistant Microorganisms and Cancer Cells

Fighting resistance to antibiotics and chemotherapeutics has brought bioactive peptides to the fore. Peptaibols are short α-aminoisobutyric acid-containing peptides produced by Trichoderma species. Here, we studied the production of peptaibols by Trichoderma atroviride O1 and evaluated their antibacterial and anticancer activity against drug-sensitive and multidrug-resistant bacterium and cancer cell lines. This was substantiated by an analysis of the activity of the peptaibol synthetase-encoding gene. Atroviridins, 20-residue peptaibols were detected using MALDI-TOF mass spectrometry. Gram-positive bacteria were susceptible to peptaibol-containing extracts of T. atroviride O1. A synergic effect of extract constituents was possible, and the biolo-gical activity of extracts was pronounced in/after the peak of peptaibol synthetase activity. The growth of methicillin-resistant Staphylococcus aureus was reduced to just under 10% compared to the control. The effect of peptaibol-containing extracts was strongly modulated by the lipoteichoic acid and only slightly by the horse blood serum present in the cultivation medium. Peptaibol-containing extracts affected the proliferation of human breast cancer and human ovarian cancer cell lines in a 2D model, including the multidrug-resistant sublines. The peptaibols influenced the size and compactness of the cell lines in a 3D model. Our findings indicate the molecular basis of peptaibol production in T. atroviride O1 and the potential of its peptaibol-containing extracts as antimicrobial/anticancer agents.     

An Efficient Method for the In Vitro Production of Azol(in)e‐Based Cyclic Peptides

Heterocycle‐containing cyclic peptides are promising scaffolds for the pharmaceutical industry but their chemical synthesis is very challenging. A new universal method has been devised to prepare these compounds by using a set of engineered marine‐derived enzymes and substrates obtained from a family of ribosomally produced and post‐translationally modified peptides called the cyanobactins. The substrate precursor peptide is engineered to have a non‐native protease cleavage site that can be rapidly cleaved. The other enzymes used are heterocyclases that convert Cys or Cys/Ser/Thr into their corresponding azolines. A macrocycle is formed using a macrocyclase enzyme, followed by oxidation of the azolines to azoles with a specific oxidase. The work is exemplified by the production of 17 macrocycles containing 6–9 residues representing 11 out of the 20 canonical amino acids.     

Emerging Methods and Design Principles for Cell‐Penetrant Peptides

Biomolecules such as antibodies, proteins, and peptides are important tools for chemical biology and leads for drug development. They have been used to inhibit a variety of extracellular proteins, but accessing intracellular proteins has been much more challenging. In this review, we discuss diverse chemical approaches that have yielded cell‐penetrant peptides and identify three distinct strategies: masking backbone amides, guanidinium group patterning, and amphipathic patterning. We summarize a growing number of large data sets, which are starting to reveal more specific design guidelines for each strategy. We also discuss advantages and disadvantages of current methods for quantifying cell penetration. Finally, we provide an overview of best‐odds approaches for applying these new methods and design principles to optimize cytosolic penetration for a given bioactive peptide.     

Combining Topology and Sequence Design for the Discovery of Potent Antimicrobial Peptide Dendrimers against Multidrug‐Resistant Pseudomonas aeruginosa

Multidrug‐resistant opportunistic bacteria, such as Pseudomonas aeruginosa, represent a major public health threat. Antimicrobial peptides (AMPs) and related peptidomimetic systems offer an attractive opportunity to control these pathogens. AMP dendrimers (AMPDs) with high activity against multidrug‐resistant clinical isolates of P. aeruginosa and Acinetobacter baumannii were now identified by a systematic survey of the peptide sequences within the branches of a distinct type of third‐generation peptide dendrimers. Combined topology and peptide sequence design as illustrated here represents a new and general strategy to discover new antimicrobial agents to fight multidrug‐resistant bacterial pathogens.     

Chemical Synthesis of Human Insulin‐Like Peptide‐6

Human insulin‐like peptide‐6 (INSL‐6) belongs to the insulin superfamily and shares the distinctive disulfide bond configuration of human insulin. In this report we present the first chemical synthesis of INSL‐6 utilizing fluorenylmethyloxycarbonyl‐based (Fmoc) solid‐phase peptide chemistry and regioselective disulfide bond construction protocols. Due to the presence of an oxidation‐sensitive tryptophan residue, two new orthogonal synthetic methodologies were developed. The first method involved the identification of an additive to suppress the oxidation of tryptophan during iodine‐mediated S‐acetamidomethyl (Acm) deprotection and the second utilized iodine‐free, sulfoxide‐directed disulfide bond formation. The methodologies presented here offer an efficient synthetic route to INSL‐6 and will further improve synthetic access to other multiple‐disulfide‐containing peptides with oxidation‐sensitive residues.     

Discovery of Macrocyclic Peptides Armed with a Mechanism-Based Warhead: Isoform-Selective Inhibition of Human Deacetylase SIRT2

Designed to inhibit: By using the random nonstandard peptide integrated discovery (RaPID) system, highly potent isoform-selective inhibitors can be identified from a library of nonstandard macrocyclic peptides. These inhibitors, which contain a mechanism-based warhead residue, are active against the human deacetylase SIRT2, with IC(50) values in the low nanomolar region.     

A One‐Pot Chemically Cleavable Bis‐Linker Tether Strategy for the Synthesis of Heterodimeric Peptides

Heterodimeric peptides linked by disulfide bonds are attractive drug targets. However, their chemical assembly can be tedious, time‐consuming, and low yielding. Inspired by the cellular synthesis of pro‐insulin in which the two constituent peptide chains are expressed as a single‐chain precursor separated by a connecting C‐peptide, we have developed a novel chemically cleavable bis‐linker tether which allows the convenient assembly of two peptide chains as a single “pro”‐peptide on the same solid support. Following the peptide cleavage and post‐synthetic modifications, this bis‐linker tether can be removed in one‐step by chemical means. This method was used to synthesize a drug delivery‐cargo conjugate, TAT‐PKCi peptide, and a two‐disulfide bridged heterodimeric peptide, thionin (7‐19)‐(24‐32R), a thionin analogue. To our knowledge, this is the first report of a one‐pot chemically cleavable bis‐linker strategy for the facile synthesis of cross‐bridged two‐chain peptides.     

Peptide Nanotubes

The self‐assembly of different classes of peptide, including cyclic peptides, amyloid peptides and surfactant‐like peptides into nanotube structures is reviewed. The modes of self‐assembly are discussed. Additionally, applications in bionanotechnology and synthetic materials science are summarized.     

Perfluoroaryl Bicyclic Cell‐Penetrating Peptides for Delivery of Antisense Oligonucleotides

Exon‐skipping antisense oligonucleotides are effective treatments for genetic diseases, yet exon‐skipping activity requires that these macromolecules reach the nucleus. While cell‐penetrating peptides can improve delivery, proteolytic instability often limits efficacy. It is hypothesized that the bicyclization of arginine‐rich peptides would improve their stability and their ability to deliver oligonucleotides into the nucleus. Two methods were introduced for the synthesis of arginine‐rich bicyclic peptides using cysteine perfluoroarylation chemistry. Then, the bicyclic peptides were covalently linked to a phosphorodiamidate morpholino oligonucleotide (PMO) and assayed for exon skipping activity. The perfluoroaryl cyclic and bicyclic peptides improved PMO activity roughly 14‐fold over the unconjugated PMO. The bicyclic peptides exhibited increased proteolytic stability relative to the monocycle, demonstrating that perfluoroaryl bicyclic peptides are potent and stable delivery agents.