Mimicry of Tandem Repeat Peptides against Cell Surface Carbohydrates

Our approach to multivalent peptide construction relies on tentacle peptides, also known as a multiple antigenic peptides, which contain two and four repeats of a selected peptide. In this communication, we report the results of preliminary studies aimed at (1) the selection of short peptides against the carbohydrate, sLeX, (2) the synthesis of tentacle dimers and tetramers of the selected peptides, and (3) the determination of affinities and specificities of the peptides to several related carbohydrates by using the surface plasmon resonance (SPR) and the equilibrium dialysis techniques. Binding affinity studies, as well as assays of in vitro binding of the peptides to a sLeX-specific cell line, have shown that the tetrameric peptides bind to the cell surface sugars

An Approach To Enhance Specificity against RNA Targets Using Heteroconjugates of Aminoglycosides and Chloramphenicol (or Linezolid)

We describe the design and synthesis of new heterodimeric conjugates, which are comprised of a neomycin B (Neo) stem-binding component and a chloramphenicol (Cam) or linezolid (Lnz) loop-binding component. Some of the heterodimeric conjugates display enhanced affinities to RNA targets and that binding occurs in both stem and loop regions of the RNA. In addition, the results of foot-printing and mutation studies suggest that the enhanced binding affinity of the conjugates is RNA sequence-specific

Combining SELEX Screening and Rational Design to Develop Light-Up Fluorophore−RNA Aptamer Pairs for RNA Tagging

We report here a new small molecule fluorogen and RNA aptamer pair for RNA labeling. The small-molecule fluorogen is designed on the basis of fluorescently quenched sulforhodamine dye. The SELEX (Systematic Evolution of Ligands by EXponential enrichment) procedure and fluorescence screening in E. coli have been applied to discover the aptamer that can specifically activate the fluorogen with micromolar binding affinity. The systematic mutation and truncation study on the aptamer structure determined the minimum binding domain of the aptamer. A series of rationally modified fluorogen analogues have been made to probe the interacting groups of fluorogen with the aptamer. These results led to the design of a much improved fluorogen ASR 7 that displayed a 33-fold increase in the binding affinity for the selected aptamer in comparison to the original ASR 1 and an 88-fold increase in the fluorescence emission after the aptamer binding. This study demonstrates the value of combining in vitro SELEX and E. coli fluorescence screening with rational modifications in discovering and optimizing new fluorogen−RNA aptamer labeling pairs

α-Helical Peptide Containing <i>N</i>,<i>N</i>-Dimethyl Lysine Residues Displays Low-Nanomolar and Highly Specific Binding to RRE RNA

The combinatorial introduction of N,N-dimethyl-Lys groups into Lys-rich α-helical peptides and measuring affinities against RRE RNA were carried out. Peptide-g, in which two Lys were replaced by N,N-dimethyl-Lys at 3 and 9 positions, showed low-nanomolar affinity, which is almost the same value as Rev peptide, the natural RRE ligand. Moreover, peptide-g displays a compatible binding specificity as Rev peptide. The effects of the positions of Lys N,N-dimethylation on the specificity of RNA binding could serve as the basis of a new strategy for the design of novel agents against RNAs. The results support that nature may use N-methylation as a post-translational modification to enhance specific peptide−RNA interactions

Efficient Method for Site-Specific ¹⁸F‑Labeling of Biomolecules Using the Rapid Condensation Reaction between 2‑Cyanobenzothiazole and Cysteine

An efficient method based on a rapid condensation reaction between 2-cyanobenzothiazole (CBT) and cysteine has been developed for ¹⁸F-labeling of N-terminal cysteine-bearing peptides and proteins. An ¹⁸F-labeled dimeric cRGD ([¹⁸F]­CBTRGD₂) has been synthesized with an excellent radiochemical yield (92% based on radio-HPLC conversion, 80% decay-corrected, and isolated yield) and radiochemical purity (>99%) under mild conditions using ¹⁸F-CBT, and shown good <i>in vivo</i> tumor targeting efficiency for PET imaging. The labeling strategy was also applied to the site-specific ¹⁸F-labeling of a protein, <i>Renilla</i> lucifierase (RLuc8) with a cysteine residue at its N-terminus. The protein labeling was achieved with 12% of decay-corrected radiochemical yield and more than 99% radiochemical purity. This strategy should provide a general approach for efficient and site-specific ¹⁸F-labeling of various peptides and proteins for <i>in vivo</i> molecular imaging applications

Isolation and Barcoding of Trace Pollen-free DNA for Authentication of Honey

Adulteration and mislabeling of honey to mask its true origin have become a global concern. Pollen microscopy, the current gold standard for identifying honey’s geographical and plant origins, is laborious, requires extensive training, and fails to identify filtered honey and honey spiked with pollen from a more favorable plant to disguise its origins. We successfully isolated pollen-free DNA from filtered honey using three types of adsorbents: (i) anti-dsDNA antibodies coupled to magnetic microspheres; (ii) anion-exchange adsorbent; and (iii) ceramic hydroxyapatite. The internal transcribed spacer 2 region of the captured pollen-free DNA was polymerase chain reaction-amplified and subjected to next-generation sequencing. Using an in-house bioinformatics pipeline, initial experiments showed that anion exchange had the greatest capacity to capture trace pollen-free DNA, and it was successfully applied to isolate DNA from five honey samples. Enrichment of trace pollen-free DNA from filtered honey samples opens a new approach for identifying the true origins of honey

Table_1_Urine ALCAM, PF4 and VCAM-1 Surpass Conventional Metrics in Identifying Nephritis Disease Activity in Childhood-Onset Systemic Lupus Erythematosus.docx

ObjectivesSerial kidney biopsy for repeat evaluation and monitoring of lupus nephritis (LN) in childhood-onset Systemic Lupus Erythematosus (cSLE) remains challenging, thus non-invasive biomarkers are needed. Here, we evaluate the performance of ten urine protein markers of diverse nature including cytokines, chemokines, and adhesion molecules in distinguishing disease activity in cSLE.MethodsEighty-four pediatric patients meeting ≥4 ACR criteria for SLE were prospectively enrolled for urine assay of 10 protein markers normalized to urine creatinine, namely ALCAM, cystatin-C, hemopexin, KIM-1, MCP-1, NGAL, PF-4, Timp-1, TWEAK, and VCAM-1 by ELISA. Samples from active renal (LN) and active non-renal SLE patients were obtained prior to onset/escalation of immunosuppression. SLE disease activity was evaluated using SLEDAI-2000. 59 patients had clinically-active SLE (SLEDAI score ≥4 or having a flare), of whom 29 patients (34.5%) were classified as active renal, and 30 patients (35.7%) were active non-renal. Twenty-five healthy subjects were recruited as controls.ResultsUrine concentrations of ALCAM, KIM-1, PF4 and VCAM-1 were significantly increased in active LN patients versus active non-renal SLE, inactive SLE and healthy controls. Five urine proteins differed significantly between 2 (hemopexin, NGAL, MCP1) or 3 (Cystatin-C, TWEAK) groups only, with the highest levels detected in active LN patients. Urine ALCAM, VCAM-1, PF4 and hemopexin correlated best with total SLEDAI as well as renal-SLEDAI scores (p ConclusionUrinary ALCAM, PF4, and VCAM-1 are potential biomarkers for predicting kidney disease activity in cSLE and hold potential as surrogate markers of nephritis flares in these patients.</p

Additional file 1 of Gender-specific relationship between thigh muscle and fat mass and brain amyloid-β positivity

Additional file 1: Table S1. Demographic variables of study participants stratified by cognitive stage