Optimizing the discovery bioanalysis strategy for macrocyclic peptides.

Macrocyclic peptides (MCPs) have remained a compelling modality in drug discovery and development, with many successful marketed drugs. Their unique molecular structure and ADME properties have posed bioanalytical challenges that cannot be fully addressed with conventional small molecule LC-MRM assays. In this work, we developed and optimized a high-throughput discovery bioanalytical strategy for MCPs with 16 marketed MCP drugs.

A New Approach for Preparing Stable High-Concentration Peptide Nanoparticle Formulations.

The subcutaneous administration of therapeutic peptides would provide significant benefits to patients. However, subcutaneous injections are limited in dosing volume, potentially resulting in high peptide concentrations that can incur significant challenges with solubility limitations, high viscosity, and stability liabilities. Herein, we report on the discovery that low-shear resonant acoustic mixing can be used as a general method to prepare stable nanoparticles of a number of peptides of diverse molecular weights and structures in water without the need for extensive amounts of organic solvents or lipid excipients.

Development of Liposome Systems for Enhancing the PK Properties of Bivalent PROTACs.

Proteolysis-Targeting Chimeras (PROTACs) are a promising new technology in drug development. They have rapidly evolved in recent years, with several of them in clinical trials. While most of these advances have been associated with monovalent protein degraders, bivalent PROTACs have also entered clinical trials, although progression to market has been limited.

Expedited Evaluation of Conformational Stability-Heterogeneity Associations for Crude Polyclonal Antibodies in Response to Conjugate Vaccines.

Conjugate vaccines have been demonstrated to be a promising strategy for immunotherapeutic intervention in substance use disorder, wherein a hapten structurally similar to the target drug is conjugated to an immunogenic carrier protein. The antibodies generated following immunization with these species can provide long-lasting protection against overdose through sequestration of the abused drug in the periphery, which mitigates its ability to cross the blood-brain barrier. However, these antibodies exhibit a high degree of heterogeneity in structure.

Research on the isolation and identification of black spot disease of Rosa chinensis in Kunming, China.

Through a survey of rose diseases in the South Tropical Garden in Kunming, China, it was found that black spot was the most common and serious disease of rose cultivated in the open air there, with an incidence of more than 90%. In this study, fungus isolation was performed on leaf samples of five black spot susceptible varieties of rose from the South Tropical Garden by tissue isolation. 18 strains of fungus were initially obtained, and seven of them were finally identified to cause black spot symptoms on healthy leaves of rose after verification by Koch's rule.

ATP-Coated Dual-Functionalized Titanium(IV) IMAC Material for Simultaneous Enrichment and Separation of Glycopeptides and Phosphopeptides.

Protein glycosylation and phosphorylation are two of the most common post-translational modifications (PTMs), which play an important role in many biological processes. However, low abundance and poor ionization efficiency of phosphopeptides and glycopeptides make direct MS analysis challenging. In this study, we developed a hydrophilicity-enhanced bifunctional Ti-IMAC (IMAC: immobilized metal affinity chromatography) material with grafted adenosine triphosphate (denoted as epoxy-ATP-Ti) to enable simultaneous enrichment and separation of common N-glycopeptides, phosphopeptides, and M6P glycopeptides from tissue/cells.

Boost-DiLeu: Enhanced Isobaric ,-Dimethyl Leucine Tagging Strategy for a Comprehensive Quantitative Glycoproteomic Analysis.

Intact glycopeptide analysis has been of great interest because it can elucidate glycosylation site information and glycan structural composition at the same time. However, mass spectrometry (MS)-based glycoproteomic analysis is hindered by the low abundance and poor ionization efficiency of glycopeptides. Relatively large amounts of starting materials are needed for the enrichment, which makes the identification and quantification of intact glycopeptides from samples with limited quantity more challenging.

First report of Chaetomium pseudocochliodes causing flower rot disease of Camellia reticulata in Yunnan, China.

Camellia reticulata is the world-famous ornamental flower (Wang et al. 2021). In February 2021, the infected flowers of C.

Analysis of pancreatic extracellular matrix protein post-translational modifications electrostatic repulsion-hydrophilic interaction chromatography coupled with mass spectrometry.

The pancreas is a vital organ with digestive and endocrine roles, and diseases of the pancreas affect millions of people yearly. A better understanding of the pancreas proteome and its dynamic post-translational modifications (PTMs) is necessary to engineer higher fidelity tissue analogues for use in transplantation. The extracellular matrix (ECM) has major roles in binding and signaling essential to the viability of insulin-producing islets of Langerhans.

Dual-Functional Ti(IV)-IMAC Material Enables Simultaneous Enrichment and Separation of Diverse Glycopeptides and Phosphopeptides.

Simultaneous enrichment and fractionation of diverse proteins/peptides possessing different post-translational modifications (PTMs) from the same biological samples is highly desirable to reduce sample consumption, avoid complicated sample processing, and enable studies of potential crosstalks between different PTMs. In this work, we report a new approach to enable simultaneous enrichment and separation of glycopeptides, phosphopeptides, and mannose-6-phosphate (M6P) glycopeptides by using a dual-functional Ti(IV)-IMAC material. Moreover, we also made the separation of neutral and sialyl glycopeptides and mono- and multi-phosphopeptides possible by performing different elution processes according to the differences in their electrostatic or hydrophilic properties.

Counterion Optimization Dramatically Improves Selectivity for Phosphopeptides and Glycopeptides in Electrostatic Repulsion-Hydrophilic Interaction Chromatography.

A well-hydrated counterion can selectively and dramatically increase retention of a charged analyte in hydrophilic interaction chromatography. The effect is enhanced if the column is charged, as in electrostatic repulsion-hydrophilic interaction chromatography (ERLIC). This combination was exploited in proteomics for the isolation of peptides with certain post-translational modifications (PTMs).

Acetyl-CoA flux from the cytosol to the ER regulates engagement and quality of the secretory pathway.

Nε-lysine acetylation in the ER is an essential component of the quality control machinery. ER acetylation is ensured by a membrane transporter, AT-1/SLC33A1, which translocates cytosolic acetyl-CoA into the ER lumen, and two acetyltransferases, ATase1 and ATase2, which acetylate nascent polypeptides within the ER lumen. Dysfunctional AT-1, as caused by gene mutation or duplication events, results in severe disease phenotypes.

Integrated Label-Free and 10-Plex DiLeu Isobaric Tag Quantitative Methods for Profiling Changes in the Mouse Hypothalamic Neuropeptidome and Proteome: Assessment of the Impact of the Gut Microbiome.

Gut microbiota can regulate host physiological and pathological status through gut-brain communications or pathways. However, the impact of the gut microbiome on neuropeptides and proteins involved in regulating brain functions and behaviors is still not clearly understood. To address the problem, integrated label-free and 10-plex DiLeu isobaric tag-based quantitative methods were implemented to compare the profiling of neuropeptides and proteins in the hypothalamus of germ-free (GF)- vs conventionally raised (ConvR)-mice.

Acetyl-CoA flux regulates the proteome and acetyl-proteome to maintain intracellular metabolic crosstalk.

AT-1/SLC33A1 is a key member of the endoplasmic reticulum (ER) acetylation machinery, transporting acetyl-CoA from the cytosol into the ER lumen where acetyl-CoA serves as the acetyl-group donor for Nε-lysine acetylation. Dysfunctional ER acetylation, as caused by heterozygous or homozygous mutations as well as gene duplication events of AT-1/SLC33A1, has been linked to both developmental and degenerative diseases. Here, we investigate two models of AT-1 dysregulation and altered acetyl-CoA flux: AT-1 mice, a model of AT-1 haploinsufficiency, and AT-1 sTg mice, a model of AT-1 overexpression.

Dual-Functional Titanium(IV) Immobilized Metal Affinity Chromatography Approach for Enabling Large-Scale Profiling of Protein Mannose-6-Phosphate Glycosylation and Revealing Its Predominant Substrates.

Mannose-6-phosphate (M6P) glycosylation is an important post-translational modification (PTM) and plays a crucial role in transferring lysosomal hydrolases to lysosome, and is involved in several other biological processes. Aberrant M6P modifications have been implicated in lysosomal storage diseases and numerous other disorders including Alzheimer's disease and cancer. Research on profiling of intact M6P glycopeptides remains challenging due to its extremely low stoichiometry.

Finding the Sweet Spot in ERLIC Mobile Phase for Simultaneous Enrichment of N-Glyco and Phosphopeptides.

Simultaneous enrichment of glyco- and phosphopeptides will benefit the studies of biological processes regulated by these posttranslational modifications (PTMs). It will also reveal potential crosstalk between these two ubiquitous PTMs. Unlike custom-designed multifunctional solid phase extraction (SPE) materials, operating strong anion exchange (SAX) resin in electrostatic repulsion-hydrophilic interaction chromatography (ERLIC) mode provides a readily available strategy to analytical labs for enrichment of these PTMs for subsequent mass spectrometry (MS)-based characterization.

Omics Technologies to Understand Activation of a Biosynthetic Gene Cluster in Micromonospora sp. WMMB235: Deciphering Keyicin Biosynthesis.

DNA sequencing of a large collection of bacterial genomes reveals a wealth of orphan biosynthetic gene clusters (BGCs) with no identifiable products. BGC silencing, for those orphan clusters that are truly silent, rather than those whose products have simply evaded detection and cluster correlation, is postulated to result from transcriptional inactivation of these clusters under standard laboratory conditions. Here, we employ a multi-omics approach to demonstrate how interspecies interactions modulate the keyicin producing kyc cluster at the transcriptome level in cocultures of kyc-bearing Micromonospora sp.

Using "On/Off" (19)F NMR/Magnetic Resonance Imaging Signals to Sense Tyrosine Kinase/Phosphatase Activity in Vitro and in Cell Lysates.

Tyrosine kinase and phosphatase are two important, antagonistic enzymes in organisms. Development of noninvasive approach for sensing their activity with high spatial and temporal resolution remains challenging. Herein, we rationally designed a hydrogelator Nap-Phe-Phe(CF3)-Glu-Tyr-Ile-OH (1a) whose supramolecular hydrogel (i.

Multifunctional Bioconjugate for Cancer Cell-Targeted Theranostics.

Cancer cell-targeted imaging and drug delivery remain a challenge for precise cancer theranostics. MUC1 is a large transmembrane glycoprotein that may potentially serve as a target for cancer theranostics. Herein, using a MUC1-targeting aptamer (APT) as the "warhead", we rationally designed and constructed a hybrid nanoparticle 1-NPs-QDs-hAPT (Vehicle) that could be applied for MUC1-targeted cell uptake and imaging.