Here we present a practical and operationally straightforward method for cysteine thioarylation under mild redox-neutral conditions. The reaction is mediated by a photoactive copper complex formed in situ from a readily available copper salt and the cysteine-containing substrate.

The development of chemical methods enabling site-selective incorporation of noncanonical amino acids into peptide backbones with precise functional tailoring remains a critical challenge. Particularly compelling is the use of underexplored endogenous amino acid hotspots, such as the N (in) of tryptophan, as versatile anchors for diversification. Herein, we report a chemoselective N(sp2)-H bond activation strategy targeting native tryptophan residues within peptide frameworks, exemplified by GLP-1 (7-37), using nickel metallaphotocatalysis under postsynthetic solid-phase conditions. This selective N (in)-arylation reaction proceeds efficiently within 3 h of light irradiation in highly functionalized heterogeneous environments, employing minimal excesses of electrophile and base, alongside catalytic quantities of nickel, ligand, and photocatalyst. The method affords homogeneous peptide products with high chemoselectivity and operational simplicity. We envision that this strategy could contribute to advancing the design of the next-generation long-acting class II G protein-coupled receptor agonist therapeutics.

The discovery of useful synthetic transformations has made light-mediated catalysis, a widely employed method in chemical synthesis. Since the catalyst, light source, and substrate needed to produce a photoredox reaction are the same as those needed for photosensitization, photoredox reactions are perfect for examining biological surroundings. An attempt has been made to cover the development of future-oriented catalysts and the therapeutic use of photosensitization. New applications of photoredox catalytic techniques for investigating intricate biological environments in living cells and protein bioconjugation is also discussed.

Antibody-drug conjugates (ADCs) represent a promising class of targeted therapeutics, combining the specificity of antibodies with the potency of cytotoxic drugs to enhance therapeutic efficacy while minimizing off-target effects. The development of new chemical methods for bioconjugation is essential to generate ADCs and to optimize their stability, efficacy, and safety. Traditional conjugation methods often face challenges related to site-selectivity and heterogeneous product mixtures, highlighting the need to develop new, innovative chemical strategies. Photoredox chemistry emerges as a powerful tool in this context, enabling precise, mild, and selective modifications of peptides and proteins. By harnessing light to drive chemical transformations, photoredox techniques can facilitate the synthesis of antibody bioconjugates. This perspective will discuss the drive to develop and empower photoredox methods applied to antibody functionalization.

The advent of bioorthogonal chemistry has transformed scientific research, offering a powerful tool for selective and noninvasive labeling of (bio)molecules within complex biological environments. This innovative approach has facilitated the study of intricate cellular processes, protein dynamics, and interactions. Nevertheless, a number of challenges remain to be addressed, including the need for improved reaction kinetics, enhanced biocompatibility, and the development of a more diverse and orthogonal set of reactions. While scientists continue to search for veritable solutions, bioorthogonal chemistry remains a transformative tool with a vast potential for advancing our understanding of biology and medicine. This Perspective offers insights into reactions commonly classified as "bioorthogonal", which, however, may not always demonstrate the desired selectivity regarding the interactions between their components and the additives or catalysts used under the reaction conditions.

Platinum-based anticancer drugs (PBCs), particularly cisplatin, play a key role in over 70% of cancer treatment protocols. PBCs suffer from their strong affinity with numerous nucleophiles present in the body, leading to significant systematic toxicity and rapid drug inactivation. The cell membrane's selective and energy-dependent transport properties, inherent to its unique biological structure, offer a strategic opportunity for employing cell membranes (CMs) in the development of PBC delivery systems that repel nucleophiles. To prove this idea, we harness cancer CMs to develop a dual-package approach for sealing cisplatin in a nanoformulation that is both nucleophile-resistant and tumor-targeted without the need for synthetic materials. The dual-package process begins by conjugating cisplatin to cancer CMs, creating positively charged nanoparticles. These isolated nanoparticles are then recomplexed with cancer CMs. Our strategy, which tightly seals cisplatin within the cancer CMs, ensures that cisplatin is safely sequestered from reactive molecules in the body while simultaneously guiding it specifically to homologous tumors. The resulting nanoformulation demonstrates immune evasion and a prolonged circulation time due to the native-like identity conferred by cancer CMs. The biomimetic sealing of cisplatin within CMs prevented the transmembrane attack of nucleophiles, including not only macromolecular proteins but also small-molecule compounds such as glutathione, thereby ensuring a high level of cytotoxicity when challenged by these nucleophiles. It also displays precise targeting at homologous tumors, ensures sustained drug release, and achieves significant tumor suppression. These features together adumbrate the nanoformulation's potential as a revolutionary tool in cisplatin cancer therapy. Given the prevalence of metal ion-based drugs and their common susceptibility to nucleophile-associated issues, the strategy presented in this study may offer a widely applicable solution to developing nucleophile-resistant metal-ion-based medications.

In spite of being the second-lowest abundant proteinogenic amino acid, approximately 90% of proteins contain at least one tryptophan residue. Hence, the chemoselective functionalization of tryptophan residue can provide access to site-selective bioconjugation of almost all known proteins. With the increase in the utility of bioconjugated proteins and peptides as drugs and therapeutic agents, the development of smart protocols to fabricate and modulate biomolecules has flourished. This review provides a comprehensive summary of the latest advances in tryptophan-specific modification and diversification of peptides and proteins that exhibit significant applications in proteomics, protein engineering, living cell imaging, drug discovery, etc. The article also highlights literature gaps and new opportunities for the sake of future innovation in the field.

Viscoelastic heterogeneity of matrices plays a pivotal role in cancer cell spreading, migration, and metastasis. However, the creation of viscoelastic platforms with spatial-temporal regulation is hindered by cytotoxicity and short regulation durations. Our research presents a dual mechanism for stress relaxation regulation- both intrinsic and responsive- by incorporating Schiff base bonds and a visible light-responsive thiuram disulfide (TDS) moiety into the hydrogel. Modifying base bonds facilitates a broad spectrum of intrinsic stress relaxation times. At the same time, incorporating the visible light-responsive TDS moiety endows the hydrogel with responsive viscoelastic properties. These properties are characterized by minimal cytotoxicity, spatial-temporal controllability, dose dependency, and reversibility. Utilizing this platform, we demonstrate that ovarian cancer cells exhibit contrasting behaviors in contraction and spreading when subjected to dynamic stress relaxation changes over various time periods. Additionally, we observed a "memory effect" in the cell's response to alterations in stress relaxation time. We can spatially direct cell migration through viscoelastic heterogeneity, achieved via photopatterning substrates and laser spots. This innovative approach provides a means to regulate the viscoelasticity of hydrogels across a wide range of timescales, thereby opening avenues for more advanced studies into how cells interpret and respond to spatiotemporal viscoelastic signals.

The glycosylation of peptides and proteins can significantly impact their intrinsic properties, such as conformation, stability, antigenicity, and immunogenicity. Current methods for preparing N-linked glycopeptides typically rely on amide bond formation, which can be limited by the presence of reactive functional groups like acids and amines. Late-stage functionalization of peptides offers a promising approach to obtaining N-linked glycopeptides. In this study, we demonstrate the preparation of N-linked glycopeptides through a photoredox-catalyzed site-selective Giese addition between N-glycosyl oxamic acid and peptides containing dehydroalanine (Dha) under visible light conditions. Unlike traditional methods that rely on the coupling of aspartic acid and glycosylamine, this approach utilizes the conjugation of N-glycosylated carbamoyl radicals with Dha, facilitating the straightforward modification of complex peptides.

The chemical synthesis of unnatural amino acids (UAA) is a key strategy for preparing designed peptides, including pharmaceutically active compounds. Alterations of existing amino acid residues such as dehydroalanine (Dha) are particularly important since selected positions can be addressed without the necessity of a complete de novo synthesis. The intriguing UAA 4,5-dihydroxynorvaline (Dnv) is found in a variety of naturally occurring peptides and biologically active compounds. However, no method is currently available to modify an existing peptide with this residue. We report the use of flavin catalysts and visible light irradiation for this challenge, which serves as a versatile strategy for converting Dha into Dnv. Our study shows that excited flavins are competent hydrogen atom abstraction catalysts for ethers and acetals, which allows masked 1,2-dihydroxyethylene functionalization from 2,2-dimethyl-1,3-dioxolane. The masked diol was successfully coupled to Dha residues, and a series of Dnv-containing products is reported. A mild and orthogonal protocol for deprotection of the acetal group was also identified, allowing free Dnv-modified peptides to be obtained. This method provides a straightforward strategy for Dnv functionalization, which is envisioned to be crucial for accessing natural products and synthetic analogues with pharmaceutical activity.

The structure of the novel photoactive nickel species was simulated by density functional theory (DFT)/time-dependent density functional theory (TD-DFT) calculations. The application of the simplified photoactive nickel catalyst was demonstrated in a photoinduced nickel-catalyzed three-component arylsulfonation of 1,6-enynes. This reaction was autopromoted and proceeded in the absence of an additional photocatalyst. This methodology exhibited mild conditions, a broad substrate scope, and high efficiency.

Selective C-H activation on complex biological macromolecules is a key goal in the field of organic chemistry. It requires thermodynamically challenging chemical transformations to be delivered under mild, aqueous conditions. 5-Methylcytosine (5mC) is a fundamentally important epigenetic modification in DNA that has major implications for biology and has emerged as a vital biomarker. Selective functionalisation of 5mC would enable new chemical approaches to tag, detect and map DNA methylation to enhance the study and exploitation of this epigenetic feature. We demonstrate the first example of direct and selective chemical oxidation of 5mC to 5-formylcytosine (5fC) in DNA, employing a photocatalytic system. This transformation was used to selectively tag 5mC. We also provide proof-of-concept for deploying this chemistry for single-base resolution sequencing of 5mC and genetic bases adenine (A), cytosine (C), guanine (G), thymine (T) in DNA on a next-generation sequencing system. This work exemplifies how photocatalysis has the potential to transform the analysis of DNA.

Highly spatiotemporal-resolved photomodulation demonstrates promise for investigating key biological events in vivo and in vitro, such as cell signaling pathways, neuromodulation, and tumor treatment without side effects. However, enhancing the performance of photomodulation tools remains challenging due to the limitations of the physicochemical properties of the photoactive molecules. Here, a compact, stable intramolecular π-π stacking conformation forming between the target molecule (naproxen) and the perylene-based photoremovable protecting group is discovered to confine the motion of the photolabile bond and then enhance the photocleavage quantum yield. In conjunction with a red-absorbing photosensitizer, the photocleavage wavelength is extended to the red region via triplet-triplet annihilation. In particular, the triplet lifetime of the prodrug can be extended via the linked steric hindrance to improve the conversion yield via TTA. Using the new photomodulation tool, it is precisely photoreleased cyclooxygenase-2 inhibitors for tumor vascular growth suppression in vivo. In combination with cisplatin, over 90% efficient inhibition of malignant breast tumors is observed via the synergistic tumor treatment strategy. These findings provide a new concept for the rational design of efficient photocleavage and have implications for photomodulating cell signaling pathways in tumor therapy, as well as laying a solid foundation for the development of phototherapeutic approaches.

Given the severe loss of species richness across diverse ecosystems, there is an urgent need to assess and monitor biodiversity on a global scale. The analysis of environmental DNA (eDNA), referring to any DNA extracted from environmental samples and subsequently sequenced, is a promising method for performing such biodiversity related studies. However, a comprehensive understanding of the factors that drive distinct eDNA degradation rates under different environmental conditions is currently missing, which limits the spatiotemporal interpretations that are possible from the eDNA-based detection of species. Here, we explore what role photochemistry may play in the fate of eDNA in aquatic ecosystems. Since few eDNA photodegradation studies have been performed, we extrapolate measured photochemical degradation dynamics from dissolved organic matter (DOM) and cellular DNA to what is expected for eDNA. Our findings show that photochemistry may dominate eDNA degradation under certain environmental conditions (e.g., DOM-rich waters with no light-limitation) and that photochemical alteration of eDNA may impact microbial respiration rates and the quantitative polymerase chain reaction (qPCR)-based detection of eDNA. We therefore encourage future studies to analyze the impact of photochemistry on eDNA degradation and provide suggested research directions that could help improve the accuracy of spatiotemporal inferences from eDNA analyses.

Our study demonstrates the exceptional efficiency of 1-methyl-4-arylurazole (MAUra) for tyrosine labeling, optimized with laccase under mild conditions, achieving a high efficiency (kcat/Km = 7.88 × 104 M-1 s-1) with minimal oxidative side reactions and selective labeling of highly exposed tyrosine sites on proteins.

The field of bioorthogonal chemistry has revolutionized our ability to interrogate and manipulate biological systems at the molecular level. However, the range of chemical reactions that can operate efficiently in biological environments without interfering with the native cellular machinery, remains limited. In this context, the rapidly growing area of photocatalysis offers a promising avenue for developing new type of bioorthogonal tools. The inherent mildness, tunability, chemoselectivity, and external controllability of photocatalytic transformations make them particularly well-suited for applications in biological and living systems. This minireview summarizes recent advances in bioorthogonal photocatalytic technologies, with a particular focus on their potential to enable the selective generation of designed products within biologically relevant or living settings.

This study reports the reversible solubility switching of a polymer triggered by non-phototoxic visible light. A photochromic polymerizable azobenzene monomer with four methoxy groups at the ortho-position (mAzoA) was synthesized, exhibiting reversible photoisomerization between trans- and cis-states using green (546 nm) and blue light (436 nm). Free radical copolymerization of hydrophilic dimethylacrylamide (DMAAm) with mAzoA produced a light-responsive random copolymer (P(mAzoA-r-DMAAm)) that shows a reversible photochromic reaction to visible light. Optimizing mAzoA content resulted in P(mAzoA10.7-r-DMAAm)3.0 kDa exhibiting LCST-type phase separation in PBS (pH 7.4) with trans- and cis-states at 39.2 °C and 32.9 °C, respectively. The bistable temperature range of 6.3 °C covers 37 °C, suitable for mammalian cell culture. Reversible solubility changes were demonstrated under alternating green and blue light at 37 °C. 1H NMR indicated significant retardation of thermal relaxation from cis- to trans-states, preventing undesired thermal mechanical degradation. Madin Darby Canine Kidney (MDCK) cells adhered to the P(mAzoA-r-DMAAm) hydrogel, confirming its non-cytotoxicity and potential for biocompatible interfaces. This principle is useful for developing hydrogels that can reversibly stimulate cells mechanically or chemically in response to visible light.

Post-translational modifications on the histone H3 tail regulate chromatin structure, impact epigenetics, and hence the gene expressions. Current chemical modulation tools, such as unnatural amino acid incorporation, protein splicing, and sortase-based editing, have allowed for the modification of histones with various PTMs in cellular contexts, but are not applicable for editing native chromatin. The use of small organic molecules to manipulate histone-modifying enzymes alters endogenous histone PTMs but lacks precise temporal and spatial control. To date, there has been no achievement in modulating histone methylation in living cells with spatiotemporal resolution. In this study, a new method is presented for temporally manipulating histone dimethylation H3K9me2 using a photo-responsive inhibitor that specifically targets the methyltransferase G9a on demand. The photo-caged molecule is stable under physiological conditions and cellular environments, but rapidly activated upon exposure to light, releasing the bioactive component that can immediately inhibit the catalytic ability of the G9a in vitro. Besides, this masked compound could also efficiently reactivate the inhibition of methyltransferase activity in living cells, subsequently suppress H3K9me2, a mark that regulates various chromatin functions. Therefore, the chemical system will be a valuable tool for manipulating the epigenome for therapeutic purposes and furthering the understanding of epigenetic mechanisms.

Domino cyclization of oxime esters and NH4SCN facilitated by photoredox and copper cocatalysis has been established. Various structurally diverse fully substituted 2-aminothiazoles have been obtained in good yields at room temperature. It is featured by mild conditions, favorable functional group tolerance, and wide substrate scope. The present reaction is amenable to gram-scale synthesis, which is expected to find potential applications in organic synthesis and drug discovery. A plausible reaction mechanism is proposed.

Sulfilimines, a privileged class of -S(iv)[double bond, length as m-dash]N- functional groups found in nature, have been exploited as valuable building blocks in organic synthesis and as pharmacophores in drug discovery, and have aroused significant interest in the chemical community. Nevertheless, strategies for late-stage introduction of sulfilimines into peptides and proteins have still met with limited success. Herein, we have developed a method of introducing biological sulfilimine fragments into peptides by an intermolecular sulfur atom transfer cascade reaction, utilizing hydroxylamine condensed with the acid moieties of peptides and varied diaryl disulfides. It provides a convenient, efficient, metal-free and widely applicable method for late-stage modification and functionalization of peptides at their acid sites both in the homogeneous phase and on-resins in SPPS. Moreover, the modified peptides with sulfilimines have been demonstrated as cleavable linkers for peptide conjugates under reducible conditions, providing unique opportunities in peptide therapeutics development and drug discovery.

Photoclick reactions combine the advantages offered by light-driven processes, that is, non-invasive and high spatiotemporal control, with classical click chemistry and have found applications ranging from surface functionalization, polymer conjugation, photocrosslinking, protein labelling and bioimaging. Despite these advances, most photoclick reactions typically require near-ultraviolet (UV) and mid-UV light to proceed. UV light can trigger undesirable responses, including cellular apoptosis, and therefore, visible and near-infrared light-induced photoclick reaction systems are highly desirable. Shifting to a longer wavelength can also reduce degradation of the photoclick reagents and products. Several strategies have been used to induce a bathochromic shift in the wavelength of irradiation-initiating photoclick reactions. For instance, the extension of the conjugated π-system, triplet-triplet energy transfer, multi-photon excitation, upconversion technology, photocatalytic and photoinitiation approaches, and designs involving photocages have all been used to achieve this goal. Current design strategies, recent advances and the outlook for long wavelength-driven photoclick reactions are presented.

With the versatile utility of bio-conjugated peptides and proteins in the fields of agriculture, food, cosmetics and pharmaceutical industry, the design of smart protocols to conjugate and modulate biomolecules becomes highly desirable. During this process, the most important consideration for biochemists is the retention of configurational integrity of the biomolecules. Moreover, this type of bioconjugation of peptide and protein becomes frivolous if the reaction is not performed with precise amino acid residues. Hence, chemo-selective, as well as site-selective reactions, that are biocompatible and possess an appropriate level of reactivity are necessary. Based on click chemistry, there are so many tyrosine (Y) conjugation strategies, such as sulfur-fluoride exchange (SuFEx), sulfur-triazole exchange (SuTEx), coupling with π-allyl palladium complexes, diazonium salts, diazodicarboxyamide-based reagents etc. Among these techniques, diazodicarboxyamide-based Y-conjugation, which is commonly known as the "tyrosine-click (Y-click) reaction", has met the expectations of synthetic and biochemists for the tyrosine-specific functionalization of biomolecules. Over the past one and a half decades, significant progress has been made in the classical organic synthesis approach, as well as its biochemical, photochemical, and electrochemical variants. Despite such progress and increasing importance, the Y-click reaction has not been reviewed to document variations in its methodology, applications, and broad utility. The present article aims to provide a summary of the approaches for the modulation of biomolecules at the hotspot of tyrosine residue by employing the Y-click reaction. The article also highlights its application for the mapping of proteins, imaging of living cells, and in the fields of analytical and medicinal chemistry.

The electron donor-acceptor (EDA) complexes have been extensively studied, which formed an electronically excited state, obviating the need for an exogenous photocatalyst. Herein, we report a mild and efficient strategy for photoinduced radical domino perfluoroalkylation/cyclization using N,N,N',N'-tetramethylethane-1,2-diamine (TMEDA) as an electron donor. This protocol could be well expanded to access various polycyclic quinazolinones containing perfluoroalkyl groups, exhibiting photocatalyst-free, good functional group tolerance, and environmentally friendly features.

The wavelength-by-wavelength resolved photoreactivity of two photo-caged carboxylic acids, i. e. 7-(diethylamino)-coumarin- and 3-perylene-modified substrates, is investigated via photochemical action plots. The observed wavelength-dependent reactivity of the chromophores is contrasted with their absorption profile. The photochemical action plots reveal a remarkable mismatch between the maximum reactivity and the absorbance. Through the action plot data, the study is able to uncover photochemical reactivity maxima at longer and shorter wavelengths, where the molar absorptivity of the chromophores is strongly reduced. Finally, the laser experiments are translated to light emitting diode (LED) irradiation and show efficient visible-light-induced release in a near fully wavelength-orthogonal, sequence-independent fashion (λLED1 = 405 nm, λLED2 = 505 nm) with both chromophores in the same reaction solution. The herein pioneered wavelength orthogonal release systems open an avenue for releasing two different molecular cargos with visible light in a fully orthogonal fashion.

Late-stage modification of peptides could potentially endow peptides with significant bioactivity and physicochemical properties, and thereby provide novel opportunities for peptide pharmaceutical studies. Since tryptophan (Trp) bears a unique indole ring residue and plays various critical functional roles in peptides, the modification methods for tryptophan were preliminarily developed with considerable progress via transition-metal mediated C-H activation. Herein, we report an unprecedented tertiary amine catalyzed peptide allylation via the SN2'-SN2' pathway between the N1 position of the indole ring of Trp and Morita-Baylis-Hillman (MBH) carbonates. Using this method that proceeds under mild conditions, we demonstrated an extremely broad scope of Trp-containing peptides and MBH carbonates to prepare a series of peptide conjugates and cyclic peptides. The reaction is amenable to either solid-phase (on resin) or solution-phase conditions. In addition, the modified peptides can be further conjugated with other biomolecules at Trp, providing a new handle for bioconjugation.

RNA modifications play crucial roles in regulating gene expression and cellular homeostasis. Modulating RNA modifications, particularly by targeting the enzymes responsible for their catalysis, has emerged as a promising therapeutic strategy. However, limitations, such as the lack of identified modifying enzymes and compensatory mechanisms, hinder targeted interventions. Chemical approaches independent of enzymatic activity offer an alternative strategy for RNA modification modulation. Here, we present the identification of 2-chloro-3,5-dinitrobenzoic acid as a highly effective photochemical deprenylase of i6A RNA. This method demonstrates exceptional selectivity towards i6A, converting its substituent into a "N-doped" ozonide, which upon hydrolysis releases natural adenine. We believe that this chemical approach will pave the way for a better understanding of RNA modification biology and the development of novel therapeutic modalities.

Hydrophilic and biocompatible hydrogels are widely applied as ideal scaffolds in tissue engineering. The "smart" gelation material can alter its structural, physiochemical, and functional features in answer to various endo/exogenous stimuli to better biomimic the endogenous extracellular matrix for the engineering of cells and tissues. Light irradiation owns a high spatial-temporal resolution, complete biorthogonal reactivity, and fine-tunability and can thus induce physiochemical reactions within the matrix of photoresponsive hydrogels with good precision, efficiency, and safety. Both gel structure (e.g., geometry, porosity, and dimension) and performance (like conductivity and thermogenic or mechanical properties) can hence be programmed on-demand to yield the biochemical and biophysical signals regulating the morphology, growth, motility, and phenotype of engineered cells and tissues. Here we summarize the strategies and mechanisms for encoding light-reactivity into a hydrogel and demonstrate how fantastically such responsive gels change their structure and properties with light irradiation as desired and thus improve their applications in tissue engineering including cargo delivery, dynamic three-dimensional cell culture, and tissue repair and regeneration, aiming to provide a basis for more and better translation of photoresponsive hydrogels in the clinic.

Peptide and protein postmodification have gained significant attention due to their extensive impact on biomolecule engineering and drug discovery, of which cysteine-specific modification strategies are prominent due to their inherent nucleophilicity and low abundance. Herein, the study introduces a novel approach utilizing multifunctional 5-substituted 1,2,3-triazine derivatives to achieve multifaceted bioconjugation targeting cysteine-containing peptides and proteins. On the one hand, this represents an inaugural instance of employing 1,2,3-triazine in biomolecular-specific modification within a physiological solution. On the other hand, as a powerful combination of precision modification and biorthogonality, this strategy allows for the one-pot dual-orthogonal functionalization of biomolecules utilizing the aldehyde group generated simultaneously. 1,2,3-Triazine derivatives with diverse functional groups allow conjugation to peptides or proteins, while bi-triazines enable peptide cyclization and dimerization. The examination of the stability of bi-triazines revealed their potential for reversible peptide modification. This work establishes a comprehensive platform for identifying cysteine-selective modifications, providing new avenues for peptide-based drug development, protein bioconjugation, and chemical biology research.

Hydrogel presents a three-dimensional polymer network with high water content. Over the past decade, hydrogel has developed from static material to intelligent material with controllable response. Various stimuli are involved in the formation of hydrogel network, among which photo-stimulation has attracted wide attention due to the advantages of controllable conditions, which has a good application prospect in the treatment of ophthalmic diseases. This paper reviews the application of photo-crosslink hydrogels in ophthalmology, focusing on the types of photo-crosslink hydrogels and their applications in ophthalmology, including drug delivery, tissue engineering and 3D printing. In addition, the limitations and future prospects of photo-crosslink hydrogels are also provided.

We introduce a novel and straightforward methodology for photoredox arylation of an indole scaffold using aryldiazonium salts under mild and metal-free conditions. Our approach enables the regioselective and chemoselective introduction of several aryl groups to the C(2) position of indoles and tryptophan, even in competition with other amino acids. This approach extends to the late-stage functionalization of peptides and lysozyme, heralding the unprecedented arylation of tryptophan residues in wild-type proteins and offering broad utility in chemical biology.

Unnatural amino acids, and their synthesis by the late-stage functionalization (LSF) of peptides, play a crucial role in areas such as drug design and discovery. Historically, the LSF of biomolecules has predominantly utilized traditional synthetic methodologies that exploit nucleophilic residues, such as cysteine, lysine or tyrosine. Herein, we present a photocatalytic hydroarylation process targeting the electrophilic residue dehydroalanine (Dha). This residue possesses an α,β-unsaturated moiety and can be combined with various arylthianthrenium salts, both in batch and flow reactors. Notably, the flow setup proved instrumental for efficient scale-up, paving the way for the synthesis of unnatural amino acids and peptides in substantial quantities. Our photocatalytic approach, being inherently mild, permits the diversification of peptides even when they contain sensitive functional groups. The readily available arylthianthrenium salts facilitate the seamless integration of Dha-containing peptides with a wide range of arenes, drug blueprints, and natural products, culminating in the creation of unconventional phenylalanine derivatives. The synergistic effect of the high functional group tolerance and the modular characteristic of the aryl electrophile enables efficient peptide conjugation and ligation in both batch and flow conditions.

Intracellular proteome aggregation is a ubiquitous disease hallmark with its composition associated with pathogenicity. Herein, this work reports on a cell-permeable photosensitizer (P8, Rose Bengal derivative) for selective photo induced proximity labeling and crosslinking of cellular aggregated proteome. Rose Bengal is identified out of common photosensitizer scaffolds for its unique intrinsic binding affinity to various protein aggregates driven by the hydrophobic effect. Further acetylation permeabilizes Rose Bengal to selectively image, label, and crosslink aggregated proteome in live stressed cells. A combination of photo-chemical, tandem mass spectrometry, and protein biochemistry characterizations reveals the complexity in photosensitizing pathways (both Type I & II), modification sites and labeling mechanisms. The diverse labeling sites and reaction types result in highly effective enrichment and identification of aggregated proteome. Finally, aggregated proteomics and interaction analyses thereby reveal extensive entangling of proteostasis network components mediated by HSP70 chaperone (HSPA1B) and active participation of autophagy pathway in combating proteasome inhibition. Overall, this work exemplifies the first photo induced proximity labeling and crosslinking method (namely AggID) to profile intracellular aggregated proteome and analyze its interactions.

We herein report a method for site-selective photo-crosslinking of a DNA duplex. A stilbene pair was introduced into a DNA duplex and a ruthenium complex was conjugated with a triplex-forming oligonucleotide. We demonstrated that [2+2] photocycloaddition of the stilbene pair occurred upon irradiation with visible light when the ruthenium complex was in close proximity due to triplex formation. No reaction occurred when the ruthenium complex was not in proximity to the stilbene pair. The wavelength of visible light used was of lower energy than the wavelength of UV light necessary for direct excitation of stilbene. Quantum chemical calculation indicated that ruthenium complex catalyzed the photocycloaddition via triplet-triplet energy transfer. Site selectivity of this photo-crosslinking system was evaluated using a DNA duplex bearing two stilbene pairs as a substrate; we showed that the site of crosslinking was precisely regulated by the sequence of the oligonucleotide linked to the ruthenium complex. Since this method does not require orthogonal photoresponsive molecules, it will be useful in construction of complex photoresponsive DNA circuits, nanodevices and biological tools.

Photon upconversion is a method for harnessing high-energy excited states from low-energy photons. Such photons, particularly in the red and near-infrared wavelength ranges, can penetrate tissue deeply and undergo less competitive absorption in coloured reaction media, enhancing the efficiency of large-scale reactions and in vivo phototherapy. Among various upconversion methodologies, the organic-based triplet-triplet annihilation upconversion (TTA-UC) stands out - demonstrating high upconversion efficiencies, requiring low excitation power densities and featuring tunable absorption and emission wavelengths. These factors contribute to improved photochemical reactions for fields such as photoredox catalysis, photoactivation, 3D printing and immunotherapy. In this Review, we explore concepts and design principles of organic TTA-UC-mediated photochemical reactions, highlighting notable advancements in the field, as well as identify challenges and propose potential solutions. This Review sheds light on the potential of organic TTA-UC to advance beyond the traditional photochemical reactions and paves the way for research in various fields and clinical applications.

The biological and biomechanical functions of cartilage, bone and osteochondral tissue are naturally orchestrated by a complex crosstalk between zonally dependent cells and extracellular matrix components. In fact, this crosstalk involves biomechanical signals and the release of biochemical cues that direct cell fate and regulate tissue morphogenesis and remodelling in vivo. Three-dimensional bioprinting introduced a paradigm shift in tissue engineering and regenerative medicine, since it allows to mimic native tissue anisotropy introducing compositional and architectural gradients. Moreover, the growing synergy between bioprinting and drug delivery may enable to replicate cell/extracellular matrix reciprocity and dynamics by the careful control of the spatial and temporal patterning of bioactive cues. Although significant advances have been made in this direction, unmet challenges and open research questions persist. These include, among others, the optimization of scaffold zonality and architectural features; the preservation of the bioactivity of loaded active molecules, as well as their spatio-temporal release; the in vitro scaffold maturation prior to implantation; the pros and cons of each animal model and the graft-defect mismatch; and the in vivo non-invasive monitoring of new tissue formation. This work critically reviews these aspects and reveals the state of the art of using three-dimensional bioprinting, and its synergy with drug delivery technologies, to pattern the distribution of cells and/or active molecules in cartilage, bone and osteochondral engineered tissues. Most notably, this work focuses on approaches, technologies and biomaterials that are currently under in vivo investigations, as these give important insights on scaffold performance at the implantation site and its interaction/integration with surrounding tissues.

Chemically modified nucleosi(ti)des and functional oligonucleotides (ONs, including therapeutic oligonucleotides, aptamer, nuclease, etc.) have been identified playing an essential role in the areas of medicinal chemistry, chemical biology, biotechnology, and nanotechnology. Introduction of functional groups into the nucleobases of ONs mostly relies on the laborious de novo chemical synthesis. Due to the importance of nucleosides modification and aforementioned limitations of functionalizing ONs, herein, we describe a highly efficient site-selective alkylation at the C8-position of guanines in guanosine (together with its analogues), GMP, GDP, and GTP, as well as late-stage functionalization of dinucleotides and single-strand ONs (including ssDNA and RNA) through photo-mediated Minisci reaction. Addition of catechol to assist the formation of alkyl radicals via in situ generated boronic acid catechol ester derivatives (BACED) markedly enhances the yields especially for the reaction of less stable primary alkyl radicals, and is the key to success for the post-synthetic alkylation of ONs. This method features excellent chemoselectivity, no necessity for pre-protection, wide range of substrate scope, various free radical precursors, and little strand lesion. Downstream applications in disease treatment and diagnosis, or as biochemical probes to study biological processes after linking with suitable fluorescent compounds are expected.

The chemical modification of biopolymers like peptides and proteins is a key technology to access vaccines and pharmaceuticals. Similarly, the tunable derivatization of individual amino acids is important as they are key building blocks of biomolecules, bioactive natural products, synthetic polymers, and innovative materials. The high diversity of functional groups present in amino acid-based molecules represents a significant challenge for their selective derivatization Recently, visible light-mediated transformations have emerged as a powerful strategy for achieving chemoselective biomolecule modification. This technique offers numerous advantages over other methods, including a higher selectivity, mild reaction conditions and high functional-group tolerance. This review provides an overview of the most recent methods covering the photoinduced modification for single amino acids and site-selective functionalization in peptides and proteins under mild and even biocompatible conditions. Future challenges and perspectives are discussed beyond the diverse types of photocatalytic transformations that are currently available.

An efficient method for the late-stage selective O-fluoroalkylation of tyrosine residues with a stable yet highly reactive fluoroalkylating reagent, 3,3-difluoroallyl sulfonium salts (DFASs), has been developed. The reaction proceeds in a mild basic aqueous buffer (pH = 11.6) with high efficiency, high biocompatibility, and excellent regio- and chemoselectivity. Various oligopeptides and phenol-containing bioactive molecules, including carbohydrates and nucleosides, could be selectively O-fluoroalkylated. The added vinyl and other functional groups from DFASs can be valuable linkers for successive modification, significantly expanding the chemical space for further bioconjugation. The synthetic utility of this protocol has been demonstrated by the fluorescently labeled anti-cancer drug and the synthesis of O-link type 1,4,7,10-tetraazacyclododecane-N,N',N,N'-tetraacetic acid-tyrosine3-octreotate (DOTA-TATE), showing the prospect of the method in medicinal chemistry and chemical biology.

α,α-Disubstituted α-amino acids (α-AAs) have improved properties compared to other types of amino acids. They serve as modifiers of peptide conformation and as precursors of bioactive compounds. Therefore, it has been a long-standing goal to construct this highly valuable scaffold efficiently in organic synthesis and drug discovery. However, access to α,α-disubstituted α-AAs is highly challenging and largely unexplored due to their steric constraints. To overcome these, remarkable advances have been made in the last decades. Emerging strategies such as synergistic enantioselective catalysis, visible-light-mediated photocatalysis, metal-free methodologies and CO2 fixation offer new avenues to access the challenging synthesis of α,α-disubstituted α-AAs and continuously bring additional contributions to this field. This review article aims to provide an overview of the recent advancements since 2015 and discuss existing challenges for the synthesis of α,α-disubstituted α-AAs and their derivatives.

Enabling control over the bioactivity of proteins with light, along with the principles of photopharmacology, has the potential to generate safe and targeted medical treatments. Installing light sensitivity in a protein can be achieved through its covalent modification with a molecular photoswitch. The general challenge in this approach is the need for the use of low energy visible light for the regulation of bioactivity. In this study, we report visible light control over the cytolytic activity of a protein. A water-soluble visible-light-operated tetra-ortho-fluoro-azobenzene photoswitch was synthesized by utilizing the nucleophilic aromatic substitution reaction for installing a solubilizing sulfonate group onto the electron-poor photoswitch structure. The azobenzene was attached to two cysteine mutants of the pore-forming protein fragaceatoxin C (FraC), and their respective activities were evaluated on red blood cells. For both mutants, the green-light-irradiated sample, containing predominantly the cis-azobenzene isomer, was more active compared to the blue-light-irradiated sample. Ultimately, the same modulation of the cytolytic activity pattern was observed toward a hypopharyngeal squamous cell carcinoma. These results constitute the first case of using low energy visible light to control the biological activity of a toxic protein.

Selective chemical reactions at precise amino acid residues of peptides and proteins have become an exploding field of research in the last few decades. With the emerging utility of bioconjugated peptides and proteins as drug leads and therapeutic agents, the design of smart protocols to modulate and conjugate biomolecules has become necessary. During this modification, the most important concern of biochemists is to keep intact the structural integrity of the biomolecules. Hence, a soft and selective biocompatible reaction environment is necessary. Electrochemistry, a mild and elegant tunable reaction platform to synthesize complex molecules while avoiding harsh and toxic chemicals, can provide such a reaction condition. However, this strategy is yet to be fully exploited in the field of selective modification of polypeptides. With this possibility, the use of electrochemistry as a reaction toolbox in peptide and protein chemistry is flourishing day by day. Unfortunately, there is no suitable review article summarizing the residue-specific modification of biomolecules. The present review provides a comprehensive summary of the latest manifested electrochemical approaches for the modulation of five redox-active amino acid residues, namely cysteine, tyrosine, tryptophan, histidine and methionine, found in peptides and proteins. The article also highlights the incredible potential of electrochemistry for the regio- as well as chemoselective bioconjugation strategy of biomolecules.

Biomolecule labeling in living systems is crucial for understanding biological processes and discovering therapeutic targets. A variety of labeling warheads have been developed for multiple biological applications, including proteomics, bioimaging, sequencing, and drug development. Quinone methides (QMs), a class of highly reactive Michael receptors, have recently emerged as prominent warheads for on-demand biomolecule labeling. Their highly flexible functionality and tunability allow for diverse biological applications, but remain poorly explored at present. In this regard, we designed, synthesized, and evaluated a series of new QM probes with a trifluoromethyl group at the benzyl position and substituents on the aromatic ring to manipulate their chemical properties for biomolecule labeling. The engineered QM warhead efficiently labeled proteins both in vitro and under living cell conditions, with significantly enhanced activity compared to previous QM warheads. We further analyzed the labeling efficacy with the assistance of density functional theory (DFT) calculations, which revealed that the QM generation process, rather than the reactivity of QM, contributes more predominantly to the labeling efficacy. Noteworthy, twelve nucleophilic residues on the BSA were labeled by the probe, including Cys, Asp, Glu, His, Lys, Asn, Gln, Arg, Ser, Thr, Trp and Tyr. Given their high efficiency and tunability, these new QM warheads may hold great promise for a broad range of applications, especially spatiotemporal proteomic profiling for in-depth biological studies.

Improving the retention of small-molecule-based therapeutic agents in tumors is crucial to achieve precise diagnosis and effective therapy of cancer. Herein, we propose a β-galactosidase (β-Gal)-activated and red light-induced RNA modification (GALIRM) strategy for prolonged tumor imaging. A β-Gal-activatable near-infrared (NIR) fluorescence (FL) and positron emission tomography (PET) bimodal probe 68Ga-NOTA-FCG consists of a triaaza triacetic acid chelator NOTA for 68Ga-labeling, a β-Gal-activated photosensitizer CyGal, and a singlet oxygen (1O2)-susceptible furan group for RNA modification. Studies have demonstrated that the probe emits an activated NIR FL signal upon cleavage by endogenous β-Gal overexpressed in the lysosomes, which is combined with the PET imaging signal of 68Ga allowing for highly sensitive imaging of ovarian cancer. Moreover, the capability of 68Ga-NOTA-FCG generating 1O2 under 690 nm illumination could be simultaneously unlocked, which can trigger the covalent cross-linking between furan and nucleotides of cytoplasmic RNAs. The formation of the probe-RNA conjugate can effectively prevent exocytosis and prolong retention of the probe in tumors. We thus believe that this GALIRM strategy may provide entirely new insights into long-term tumor imaging and efficient tumor treatment.

Light provides high temporal precision for neuronal modulations. Small molecules are advantageous for neuronal modulation due to their structural diversity, allowing them to suit versatile targets. However, current optochemical methods release uncaged small molecules with uniform concentrations in the irradiation area, which lack spatial specificity as counterpart optogenetic methods from genetic encoding for photosensitive proteins. Photocatalysis provides spatial specificity by generating reactive species in the proximity of photocatalysts. However, current photocatalytic methods use antibody-tagged heavy-metal photocatalysts for spatial specificity, which are unsuitable for neuronal applications. Here, we report a genetically encoded metal-free photocatalysis method for the optochemical modulation of neurons via deboronative hydroxylation. The genetically encoded photocatalysts generate doxorubicin, a mitochondrial uncoupler, and baclofen by uncaging stable organoboronate precursors. The mitochondria, nucleus, membrane, cytosol, and ER-targeted drug delivery are achieved by this method. The distinct signaling pathway dissection in a single projection is enabled by the dual optogenetic and optochemical control of synaptic transmission. The itching signaling pathway is investigated by photocatalytic uncaging under live-mice skin for the first time by visible light irradiation. The cell-type-specific release of baclofen reveals the GABABR activation on NaV1.8-expressing nociceptor terminals instead of pan peripheral sensory neurons for itch alleviation in live mice.

Bacterial infections from chronic wounds affect about 175 million people each year and are a significant clinical problem. Through the integration of photodynamic therapy (PDT) and chemotherapy, a new photosensitizer consisting of ammonium salt N,N-bis-(2-hydroxyethyl)-N-(6-(4-(10,15,20-trimesitylporphyrin-5-yl) phenoxy) hexane)-N-methanaminium bromide, TMP(+) was successfully synthesized with a total reaction yield of 10%. The novel photosensitizer consists of two parts, a porphyrin photosensitizer part and a quaternary ammonium salt part, to achieve the synergistic effect of photodynamic and chemical antibacterial activity. With the increase of TMP(+) concentration, the diameter of the PCT fiber membranes (POL/COL/TMP(+); POL, polycaprolactone; COL, collagen) gradually increased, which was caused by the charge of the quaternary ammonium salt. At the same time, the antibacterial properties were gradually improved. We finally selected the PCT 0.5% group for the antibacterial experiment, with excellent performance in fiber uniformity, hydrophobicity and biosafety. The antibacterial experiment showed that the modified porphyrin TMP(+) had a better antibacterial effect than others. In vivo chronic wound healing experiments proved that the antibacterial and anti-inflammatory effect of the PCTL group was the best, further confirmed by H&E histological analysis, immunofluorescence and immunohistochemistry mechanism experiments. This research lays the foundation for the manufacture of novel molecules that combine chemical and photodynamic strategies.