Free-Radical-Mediated Photoinduced Electron Transfer between 6-Thioguanine and Tryptophan Leading to DNA-Protein-Like Cross-Link.

The nucleobase analog 6-thioguanine (6-TG) has emerged as important immunosuppressant, anti-inflammatory, and anticancer drug in the past few decades, but its unique photosensitivity of absorbing strongly ultraviolet UVA light elicits photochemical hazards in many ways. The particularly intriguing yet unresolved question is whether the direct photoreaction of 6-TG can promote DNA-protein cross-links (DPCs) formation, which are large DNA adducts blocking DNA replication and physically impede DNA-related processes. Herein, by real-time observation of radical intermediates using time-resolved UV-vis absorption spectroscopy in conjunction with product analysis by HPLC-MS, we discover that UVA excitation of 6-TG triggers direct covalent cross-linking with tryptophan (TrpH) via an exquisite radical mechanism of electron transfer. The photoexcitation prepares the redox-active triplet 36-TG*, which initiates electron transfer with TrpH, creating TrpH•+ and 6-TG•- in the first step. The deprotonated Trp• undergoes radical-recombination with its geminate partner 6-TG•- and eliminates a H2S, leading to the cross-linking product 6-TG-Trp. The photoadduct structures (two chiral isomers and one constitutional isomer) are identified unambiguously, validating further the mechanism. These findings pinpoint the exact amino acid that is vulnerable to photo-cross-linking with 6-TG and establish a mechanistic framework for understanding mutagenic DPCs formation and developing photoprobes based on this new type of photo-cross-linking.

Target-triggered and controlled release plasmon-enhanced fluorescent AIE probe for conformational monitoring of insulin fibrillation.

In this work, we constructed a target-triggered and controlled-release plasmon-enhanced fluorescent AIE probe to realize the purpose of conformational monitoring of insulin fibrillation. We synthesized a novel water-soluble anthracene derivative, 4,4',4'',4'''-(anthracene-9,10-diylbis(ethene-2,1,1-triyl))tetrakis(N,N,N-trimethylbenzenaminium) iodide (BDVAI), with AIE properties, high biocompatibility and good self-assembly effect. Gold nanocages (AuNCs) were selected as the substrate for PEF, and the inner space of hollow AuNCs was filled with BDVAI. Thiol-modified DNA chains were bonded to the surface of AuNCs by Au-S bonds, and an insulin aptamer was combined with the sulfhydryl chain to seal the AuNCs. This PEF-AIE sensor produces different fluorescence signals when interacting with native insulin and fibrillar insulin; thus, monitoring conformational changes in insulin can be realized by detecting fluorescence intensity changes during insulin fibrillation. Based on this design, this system realized sensitive detection of fibrillar insulin with a detection limit of 23.6 pM. This AIE molecular-based PEF fluorescence enhancement system improves the optical properties of fluorescent substances, which is of great significance in improving the detection sensitivity of amyloid fibrils conformational changes and providing a reliable basis for further understanding the pathogenesis of amyloidosis.

Peptide-capped functionalized Ag/Au bimetal nanoclusters with enhanced red fluorescence for lysosome-targeted imaging of hypochlorite in living cells.

Bioimaging probes for monitoring intracellular reactive oxygen species have important implications for cell biology research. Herein, we developed peptide-capped silver/gold nanoclusters (peptide@Ag/Au NCs) for lysosome-targeted imaging of hypochlorite (ClO-). The peptide@Ag/Au NCs were synthesized via a one-pot method using peptide as both a template and a reducing agent. The fluorescence intensity and absolute quantum yield of peptide@Ag/Au NCs were much higher than those of peptide-capped gold nanoclusters and silver nanoclusters. In the presence of ClO-, the fluorescence of peptide@Ag/Au NCs was quenched, accompanied by a redshift due to ClO--induced oxidation of the peptide ligand and decreased Ag content in Ag/Au NCs. The relative fluorescence intensity F0/F had favourable linearity for ClO- concentrations in the range 0.1-100 µmol/L (R2 = 0.9954), with a detection limit (LOD) of 80 nmol/L. The lysosome-targeted peptide@Ag/Au NCs were applied to detect ClO- in lysosomes in living cells via fluorescence imaging.

Preferential Binding of π-Ligand Porphyrin Targeting 5'-5' Stacking Interface of Human Telomeric RNA G-Quadruplex Dimer.

Human telomeric RNA (TERRA) containing thousands of G-rich repeats has the propensity to form parallel-stranded G-quadruplexes. The emerging crucial roles of TERRA G-quadruplexes in RNA biology fuel increasing attention for studying anticancer ligand binding with such structures, which, however, remains scarce. Here we utilized multiple steady-state and time-resolved spectroscopy analyses in conjunction with NMR methods and investigated thoroughly the binding behavior of TMPyP4 to a TERRA G-quadruplex dimer formed by the 10-nucleotide sequence r(GGGUUAGGGU). It is clearly identified that TMPyP4 intercalates into the 5'-5' stacking interface of two G-quadruplex blocks with a binding stoichiometry of 1:1 and binding constant of 1.92 × 106 M-1. This is consistent with the unique TERRA structural features of the enlarged π-π stacking plane of the A·(G·G·G·G)·A hexad at 5'-ends of each G-quadruplex block. The preferential binding of π-ligand porphyrin to the 5'-5' stacking interface of the native TERRA G-quadruplex dimer is first ascertained by the combination of dynamics and structural characterization.

PRMT5 regulates Golgi apparatus structure through methylation of the golgin GM130.

Maintenance of the Golgi apparatus (GA) structure and function depends on Golgi matrix proteins. The posttranslational modification of Golgi proteins such as phosphorylation of members of the golgin and GRASP families is important for determining Golgi architecture. Some Golgi proteins including golgin-84 are also known to be methylated, but the function of golgin methylation remains unclear. Here, we show that the protein arginine methyltransferase 5 (PRMT5) localizes to the GA and forms complexes with several components involved in GA ribbon formation and vesicle tethering. PRMT5 interacts with the golgin GM130, and depletion of PRMT5 causes defects in Golgi ribbon formation. Furthermore, PRMT5 methylates N-terminal arginines in GM130, and such arginine methylation appears critical for GA ribbon formation. Our findings reveal a molecular mechanism by which PRMT5-dependent arginine methylation of GM130 controls the maintenance of GA architecture.