RESUMO
Molecular recognition binding sites that specifically identify a target molecule are essential for life science research, clinical diagnoses, and therapeutic development. Corona phase molecular recognition is a technique introduced to generate synthetic recognition at the surface of a nanoparticle corona, but it remains an important question whether such entities can achieve the specificity of natural enzymes and receptors. In this work, we generate and screen a library of 24 amphiphilic polymers, preselected for molecular recognition and based on functional monomers including methacrylic acid, acrylic acid, and styrene, iterating upon a poly(methacrylic acid-co-styrene) motif. When complexed to a single-walled carbon nanotube, some of the resulting corona phases demonstrate binding specificity remarkably similar to that of phosphodiesterase type 5 (PDE5), an enzyme that catalyzes the hydrolysis of secondary messenger. The corona phase binds selectively to a PDE5 inhibitor, Vardenafil, as well as its molecular variant, but not to other potential off-target inhibitors. Our work herein examines the specificity and sensitivity of polymer "mutations" to the corona phase, as well as direct competitions with the native binding PDE5. Using structure perturbation, corona surface characterization, and molecular dynamics simulations, we show that the molecular recognition is associated with the unique three-dimensional configuration of the corona phase formed at the nanotube surface. This work conclusively shows that corona phase molecular recognition can mimic key aspects of biological recognition sites and drug targets, opening up possibilities for pharmaceutical and biological applications.
Assuntos
Materiais Biomiméticos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5 , Nanotubos de Carbono/química , Sítios de Ligação , Materiais Biomiméticos/química , Materiais Biomiméticos/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/química , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/metabolismo , Simulação de Dinâmica Molecular , Dicloridrato de Vardenafila/química , Dicloridrato de Vardenafila/metabolismoRESUMO
Heterotrimeric G proteins communicate signals from activated G protein-coupled receptors to downstream effector proteins. In the phototransduction pathway responsible for vertebrate vision, the G protein-effector complex is composed of the GTP-bound transducin α subunit (GαT·GTP) and the cyclic GMP (cGMP) phosphodiesterase 6 (PDE6), which stimulates cGMP hydrolysis, leading to hyperpolarization of the photoreceptor cell. Here we report a cryo-electron microscopy (cryoEM) structure of PDE6 complexed to GTP-bound GαT. The structure reveals two GαT·GTP subunits engaging the PDE6 hetero-tetramer at both the PDE6 catalytic core and the PDEγ subunits, driving extensive rearrangements to relieve all inhibitory constraints on enzyme catalysis. Analysis of the conformational ensemble in the cryoEM data highlights the dynamic nature of the contacts between the two GαT·GTP subunits and PDE6 that supports an alternating-site catalytic mechanism.
Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/metabolismo , Transdução de Sinais , Transducina/metabolismo , Animais , Biocatálise , Domínio Catalítico , Bovinos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/química , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/ultraestrutura , Guanosina Trifosfato/química , Guanosina Trifosfato/metabolismo , Modelos Moleculares , Ligação Proteica , Domínios Proteicos , Transducina/química , Transducina/ultraestrutura , Dicloridrato de Vardenafila/química , Dicloridrato de Vardenafila/metabolismoRESUMO
Vardenafil, a remedy for erectile dysfunction, is easily modified, facilitating the creation of analogues that have been illegally added to functional foods and counterfeit medications. However, the medical profile of these analogues, including their safety, efficacy, safe drug combinations, metabolism and excretion, has not been completely evaluated, which could cause serious health problems. In this study, two representative vardenafil analogues, pseudovardenafil and hydroxyvardenafil, were metabolized with in-vitro model (human liver microsome) and in-vivo model (rats). The metabolized samples were extracted and characterized, using liquid chromatography quadrupole-time of flight mass spectrometry (LC-Q-TOF-MS). Some imprecise interpretations were evaluated with tandem mass spectrometry (LC-Q-TOF-MS/MS) for mass fragmentation analysis. A total of 11 metabolites of pseudovardenafil and 13 metabolites of hydroxyvardenafil that were identified have never been reported. These new metabolites could be usefully applied to forensic science and other metabolic fields. Furthermore, they could serve as principal references for the toxicity, danger, and side effects of unlawful vardenafil counterfeits.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas em Tandem/métodos , Dicloridrato de Vardenafila , Animais , Humanos , Masculino , Microssomos Hepáticos/metabolismo , Ratos , Ratos Sprague-Dawley , Dicloridrato de Vardenafila/análogos & derivados , Dicloridrato de Vardenafila/análise , Dicloridrato de Vardenafila/metabolismoRESUMO
Sulfoximines have gained considerable recognition as an important structural motif in drug discovery of late. In particular, the clinical kinase inhibitors for the treatment of cancer, roniciclib (pan-CDK inhibitor), BAYâ 1143572 (P-TEFb inhibitor), and AZDâ 6738 (ATR inhibitor), have recently drawn considerable attention. Whilst the interest in this underrepresented functional group in drug discovery is clearly on the rise, there remains an incomplete understanding of the medicinal-chemistry-relevant properties of sulfoximines. Herein we report the synthesis and inâ vitro characterization of a variety of sulfoximine analogues of marketed drugs and advanced clinical candidates to gain a better understanding of this neglected functional group and its potential in drug discovery.