RESUMEN
Aminopeptidases, enzymes with critical roles in human body, are emerging as vital biomarkers for metabolic processes and diseases. Aberrant aminopeptidase levels are often associated with diseases, particularly cancer. Small-molecule probes, such as fluorescent, fluorescent/photoacoustics, bioluminescent, and chemiluminescent probes, are essential tools in the study of aminopeptidases-related diseases. The fluorescent probes provide real-time insights into protein activities, offering high sensitivity in specific locations, and precise spatiotemporal results. Additionally, photoacoustic probes offer signals that are able to penetrate deeper tissues. Bioluminescent and chemiluminescent probes can enhance inâ vivo imaging abilities by reducing the background. This comprehensive review is focused on small-molecule probes that respond to four key aminopeptidases: aminopeptidase N, leucine aminopeptidase, Pyroglutamate aminopeptidase 1, and Prolyl Aminopeptidase, and their utilization in imaging tumors and afflicted regions. In this review, the design strategy of small-molecule probes, the variety of designs from previous studies, and the opportunities of future bioimaging applications are discussed, serving as a roadmap for future research, sparking innovations in aminopeptidase-responsive probe development, and enhancing our understanding of these enzymes in disease diagnostics and treatment.
Asunto(s)
Aminopeptidasas , Colorantes Fluorescentes , Humanos , Aminopeptidasas/metabolismo , Colorantes Fluorescentes/química , Colorantes Fluorescentes/síntesis química , Sondas Moleculares/química , Imagen Óptica , Animales , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/síntesis química , Neoplasias/diagnóstico por imagenRESUMEN
In this study, a series of new artificial luciferases (ALucs) was created using sequential insights on missing peptide blocks, which were revealed using the alignment of existing ALuc sequences. Through compensating for the missing peptide blocks in the alignment, 10 sibling sequences were artificially fabricated and named from ALuc55 to ALuc68. The phylogenetic analysis showed that the new ALucs formed an independent branch that was genetically isolated from other natural marine luciferases. The new ALucs successfully survived and luminesced with native coelenterazine (nCTZ) and its analogs in living mammalian cells. The results showed that the bioluminescence (BL) intensities of the ALucs were interestingly proportional to the length of the appended peptide blocks. The computational modeling revealed that the appended peptide blocks created a flexible region near the active site, potentially modulating the enzymatic activities. The new ALucs generated various colors with maximally approximately 90 nm redshifted BL spectra in orange upon reaction with the authors' previously reported 1- and 2-series coelenterazine analogs. The utilities of the new ALucs in bioassays were demonstrated through the construction of single-chain molecular strain probes and protein fragment complementation assay (PCA) probes. The success of this study can guide new insights into how we can engineer and functionalize marine luciferases to expand the toolbox of optical readouts for bioassays and molecular imaging.
Asunto(s)
Bioensayo , Sondas Moleculares , Animales , Chlorocebus aethiops , Filogenia , Células COS , Luciferasas/química , Mediciones Luminiscentes/métodos , Mamíferos/metabolismoRESUMEN
Pyroglutamate aminopeptidase (PGP) specifically cleaves the peptide bond of pyroglutamic acid linked to the N-terminal end of a polypeptide or protein. Previous studies showed that PGP was associated with several physiological processes and diseases especially those involving inflammation. Utilizing a 'caging' strategy, we designed and synthesized a bioluminescence probe (PBL) with a limit-of-detection of 3.7 * 10-4 mU/mL. In vivo imaging in a mouse model of inflammatory liver disease revealed that the probe has excellent sensitivity and selectivity and provides a powerful tool for studying the physiological and pathological processes involving PGP.
Asunto(s)
Modelos Animales de Enfermedad , Inflamación/diagnóstico por imagen , Sustancias Luminiscentes/química , Piroglutamil-Peptidasa I/análisis , Animales , Diagnóstico por Imagen , Inflamación/metabolismo , Sustancias Luminiscentes/síntesis química , Ratones , Estructura Molecular , Piroglutamil-Peptidasa I/metabolismoRESUMEN
Cysteine (Cys) is a semi-essential amino acid that exerts a vital role in numerous biological functions. A noninvasive method for in vivo imaging of cysteine could represent a valuable tool for research cysteine and its complex contributions in living organisms. Thus, we developed a turn-on bioluminescence probe (CBP) not only for detecting exogenous and endogenous cysteine in vitro and in vivo, but also for visualizing these cysteines in whole animal. The current applications may help shed light on the complex mechanisms of cysteine in miscellaneous physiological and pathological processes.
Asunto(s)
Cisteína/química , Colorantes Fluorescentes/química , Animales , Permeabilidad de la Membrana Celular , Humanos , Límite de Detección , Mediciones Luminiscentes , Maleatos/química , Ratones , Modelos Animales , Imagen ÓpticaRESUMEN
Iron plays an essential role in biological system. An approach for in vivo imaging of this metal ion is needed to investigate its complex contributions to physiological and pathological processes. Herein, we present a bioluminescent probe FP-1 as a powerful tool for targeting Fe2+ detection in vitro and in vivo. The turn-on sensing scheme is based on the caged strategy of luciferin-luciferase system. FP-1 not only can detect accumulations of exogenous Fe2+ in living animal, but also has the capability of monitoring labile endogenous Fe2+ levels in animal model of sepsis. Implementation of this technique provides a valuable opportunity for understanding underlying mechanisms of Fe2+ in biological processes and disease states.
Asunto(s)
Luciferina de Luciérnaga/análogos & derivados , Hierro/análisis , Sustancias Luminiscentes/química , Piridinas/química , Sepsis/metabolismo , Animales , Modelos Animales de Enfermedad , Luciérnagas/enzimología , Luciferina de Luciérnaga/síntesis química , Hierro/metabolismo , Límite de Detección , Luciferasas de Luciérnaga/química , Sustancias Luminiscentes/síntesis química , Mediciones Luminiscentes/métodos , Masculino , Ratones Transgénicos , Piridinas/síntesis químicaRESUMEN
To develop a strategy for visualizing palladium species in the biological system, several turn-on bioluminescent probes were designed and synthesized by using a Pd-induced reductive reaction herein. Such probes exhibited high sensitivity (detection limit: 0.5⯵M) and excellent selectivity toward Pd2+in vitro. In particular, probe 2 was identified as a viable molecule with the capability of visualizing the fluctuations of level and distribution of Pd2+ in living cells and animals, which provides a valuable tool for tracing Pd2+ in biological system.
Asunto(s)
Luciferasas/metabolismo , Sustancias Luminiscentes/química , Imagen Óptica/métodos , Paladio/análisis , Animales , Línea Celular , Supervivencia Celular , Límite de Detección , Mediciones Luminiscentes , Ratones , Paladio/químicaRESUMEN
Iron (Fe) is the most abundant redox-active metal ion in the human body, and its redox-active inter-convertible multiple oxidation states contributes to numerous essential biological processes. Moreover, iron overload can potentially cause cellular damage and death, as wel as numerous diseases through the aberrant production of highly reactive oxidative species (hROS). Protein-free or weakly-protein-bound Fe ions play a pivotal role as catalytic reactants of the Fenton reaction. In this reaction, hROS, such as hydroxyl radicals and high valent-iron-oxo species, are generated by a reaction between hydrogen peroxide and Fe(II), which is re-generated through reduction using abundant intracellular reductants, such as glutathione. This results in the catalytic evolution of hROS. Thus, selective detection of the catalytic Fe(II) in the living systems can explain both of the pathological and physiological functions of Fe(II). Written from the perspective of their modes of actions, this paper presents recent advances in the development of fluorescent and bioluminescent probes that can selectively detect catalytic Fe(II) together with their biological applications.
Asunto(s)
Colorantes Fluorescentes/química , Radical Hidroxilo/química , Iones/aislamiento & purificación , Hierro/aislamiento & purificación , Catálisis , Compuestos Ferrosos/química , Compuestos Ferrosos/metabolismo , Humanos , Peróxido de Hidrógeno/química , Peróxido de Hidrógeno/metabolismo , Radical Hidroxilo/metabolismo , Iones/química , Iones/metabolismo , Hierro/química , Hierro/metabolismo , Oxidación-Reducción , Especies Reactivas de Oxígeno/químicaRESUMEN
A highly sensitive and selective bioluminescent probe for hydrazine (BPH) was designed, synthesized and evaluated for detection of hydrazine in vitro and in vivo. BPH was designed to include a specific recognition group (acetyl) of hydrazine at an appropriate modification site of the optical reporter hydroxyluciferin (D-luciferin), which showed excellent performance both in selectivity and sensitivity to hydrazine. The results showed that the bioluminescent probe BPH developed in this study is an innovative and widely applicable tool for detecting hydrazine in complex natural environment or in animals.