<i>C</i>‑Terminal Acetylene Derivatized Peptides <i>via</i> Silyl-Based Alkyne Immobilization

A new Silyl-based Alkyne Modifying (SAM)-linker for the synthesis of <i>C</i>-terminal acetylene-derivatized peptides is reported. The broad scope of this SAM2-linker is illustrated by manual synthesis of peptides that are side-chain protected, fully deprotected, and disulfide-bridged. Synthesis of a 14-meric (KLAKLAK)₂ derivative by microwave-assisted automated SPPS and a one-pot cleavage click procedure yielding protected 1,2,3-triazole peptide conjugates are also described

Efficient Reagent-Saving Method for the N‑Terminal Labeling of Bioactive Peptides with Organometallic Carboxylic Acids by Solid-Phase Synthesis

Labeling of biomolecules with organometallic moieties holds great promise as a tool for chemical biology and for the investigation of biochemical signaling pathways. Herein, we report a robust and reproducible synthetic strategy for the synthesis of ruthenocenecarboxylic acid, giving the acid in 53% overall yield. This organometallic label was conjugated via solid-phase peptide synthesis in near-quantitative yield to a number of different biologically active peptides, using only 1 equiv of the acid and coupling reagents, thereby avoiding wasting the precious organometallic acid. This optimized method of stoichiometric N-terminal acylation was then also successfully applied to conjugating ferrocenecarboxylic acid and a novel organometallic Re^I(CO)₃ complex, showing the generality of the synthetic procedure

Efficient Reagent-Saving Method for the N‑Terminal Labeling of Bioactive Peptides with Organometallic Carboxylic Acids by Solid-Phase Synthesis

Labeling of biomolecules with organometallic moieties holds great promise as a tool for chemical biology and for the investigation of biochemical signaling pathways. Herein, we report a robust and reproducible synthetic strategy for the synthesis of ruthenocenecarboxylic acid, giving the acid in 53% overall yield. This organometallic label was conjugated via solid-phase peptide synthesis in near-quantitative yield to a number of different biologically active peptides, using only 1 equiv of the acid and coupling reagents, thereby avoiding wasting the precious organometallic acid. This optimized method of stoichiometric N-terminal acylation was then also successfully applied to conjugating ferrocenecarboxylic acid and a novel organometallic Re^I(CO)₃ complex, showing the generality of the synthetic procedure

Nucleoapzymes: Hemin/G-Quadruplex DNAzyme–Aptamer Binding Site Conjugates with Superior Enzyme-like Catalytic Functions

A novel concept to improve the catalytic functions of nucleic acids (DNAzymes) is introduced. The method involves the conjugation of a DNA recognition sequence (aptamer) to the catalytic DNAzyme, yielding a hybrid structure termed “nucleoapzyme”. Concentrating the substrate within the “nucleoapzyme” leads to enhanced catalytic activity, displaying saturation kinetics. Different conjugation modes of the aptamer/DNAzyme units and the availability of different aptamer sequences for a substrate provide diverse means to design improved catalysts. This is exemplified with (i) The H₂O₂-mediated oxidation of dopamine to aminochrome using a series of hemin/G-quadruplex-dopamine aptamer nucleoapzymes. All nucleoapzymes reveal enhanced catalytic activities as compared to the separated DNAzyme/aptamer units, and the most active nucleoapzyme reveals a 20-fold enhanced activity. Molecular dynamics simulations provide rational assessment of the activity of the various nucleoapzymes. The hemin/G-quadruplex–aptamer nucleoapzyme also stimulates the chiroselective oxidation of l- vs d-DOPA by H₂O₂. (ii) The H₂O₂-mediated oxidation of <i>N</i>-hydroxy-l-arginine to l-citrulline by a series of hemin/G-quadruplex–arginine aptamer conjugated nucleoapzymes

Silyl-Based Alkyne-Modifying Linker for the Preparation of C‑Terminal Acetylene-Derivatized Protected Peptides

A novel linker for the synthesis of C-terminal acetylene-functionalized protected peptides is described. This SAM1 linker is applied in the manual Fmoc-based solid-phase peptide synthesis of Leu-enkephalin and in microwave-assisted automated synthesis of Maculatin 2.1, an antibacterial peptide that contains 18 amino acid residues. For the cleavage, treatment with tetramethylammonium fluoride results in protected acetylene-derivatized peptides. Alternatively, a one-pot cleavage-click procedure affords the protected 1,2,3-triazole conjugate in high yields after purification

Short Antibacterial Peptides with Significantly Reduced Hemolytic Activity can be Identified by a Systematic l‑to‑d Exchange Scan of their Amino Acid Residues

High systemic toxicity of antimicrobial peptides (AMPs) limits their clinical application to the treatment of topical infections; in parenteral systemic application of AMPs the problem of hemolysis is one of the first to be tackled. We now show that the selectivity of lipidated short synthetic AMPs can be optimized substantially by reducing their hemolytic activity without affecting their activity against methicillin resistant <i>Staphylococcus aureus</i> (MRSA). In order to identify the optimized peptides, two sets of 32 diastereomeric H-^dArg-WRWRW-^lLys­(C­(O)­C_<i>n</i>H_2<i>n</i>+1)-NH₂ (<i>n</i> = 7 or 9) peptides were prepared using a split–split procedure to perform a systematic l-to-d exchange scan on the central WRWRW-fragment. Compared to the all-l C₈-lipidated lead sequence, diastereomeric peptides had very similar antibacterial properties, but were over 30 times less hemolytic. We show that the observed hemolysis and antibacterial activity is affected by both differences in lipophilicity of the different peptides and specific combinations of l- and d-amino acid residues. This study identified several peptides that can be used as tools to precisely unravel the origin of hemolysis and thus help to design even further optimized nontoxic very active short antibacterial peptides

Tuning the Activity of a Short Arg-Trp Antimicrobial Peptide by Lipidation of a C- or N‑Terminal Lysine Side-Chain

The attachment of lipids to <i>C</i>- or <i>N</i>-terminally positioned lysine side-chain amino groups increases the activity of a short synthetic (Arg-Trp)₃ antimicrobial peptide significantly, making these peptides even active against pathogenic Gram-negative bacteria. Thus, a peptide with strong activity against <i>S. aureus</i> (1.1–2 μM) and good activity against <i>A. baumannii</i> and <i>P. aeruginosa</i> (9–18 μM) was identified. The most promising peptide causes 50% hemolysis at 285 μM and shows some selectivity against human cancer cell lines. Interestingly, the increased activity of ferrocenoylated peptides is mostly due to the lipophilicity of the organometallic fragment

The Chemoselective Reactions of Tyrosine-Containing G-Protein-Coupled Receptor Peptides with [Cp*Rh(H₂O)₃](OTf)₂, Including 2D NMR Structures and the Biological Consequences

The bioconjugation of organometallic complexes with peptides has proven to be a novel approach for drug discovery. We report the facile and chemoselective reaction of tyrosine-containing G-protein-coupled receptor (GPCR) peptides with [Cp*Rh­(H₂O)₃]­(OTf)₂, in water, at room temperature, and at pH 5–6. We have focused on three important GPCR peptides; namely, [Tyr¹]-leu-enkephalin, [Tyr⁴]-neurotensin­(8-13), and [Tyr³]-octreotide, each of which has a different position for the tyrosine residue, together with competing functionalities. Importantly, all other functional groups present, i.e., amino, carboxyl, disulfide, phenyl, and indole, were not prominent sites of reactivity by the Cp*Rh tris aqua complex. Furthermore, the influence of the Cp*Rh moiety on the structure of [Tyr³]-octreotide was characterized by 2D NMR, resulting in the first representative structure of an organometallic-peptide complex. The biological consequences of these Cp*Rh-peptide complexes, with respect to GPCR binding and growth inhibition of MCF7 and HT29 cancer cells, will be presented for [(η⁶-Cp*Rh-Tyr¹)-leu-enkephalin]­(OTf)₂ and [(η⁶-Cp*Rh-Tyr³)-octreotide]­(OTf)₂

Trivalent Ultrashort Lipopeptides are Potent pH Dependent Antifungal Agents

The activity of antimicrobial peptides (AMPs) that contain a large proportion of histidine residues (p<i>K</i>_a ∼ 6) depends on the physiological pH environment. Advantages of these AMPs include high activity in slightly acidic areas of the human body and relatively low toxicity in other areas. Also, many AMPs are highly active in a multivalent form, but this often increases toxicity. Here we designed pH dependent amphiphilic compounds consisting of multiple ultrashort histidine lipopeptides on a triazacyclophane scaffold, which showed high activity toward Aspergillus fumigatus and Cryptococcus neoformans at acidic pH, yet remained nontoxic. In vivo, treatment with a myristic acid conjugated trivalent histidine–histidine dipeptide resulted in 55% survival of mice (<i>n</i> = 9) in an otherwise lethal murine lung Aspergillus infection model. Fungal burden was assessed and showed completely sterile lungs in 80% of the mice (<i>n</i> = 5). At pH 5.5 and 7.5, differing peptide–membrane interactions and peptide nanostructures were observed. This study underscores the potential of unique AMPs to become the next generation of clinical antimicrobial therapy