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1.
Genes Dev ; 38(11-12): 528-535, 2024 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-38960718

RESUMO

As part of the efforts to understand nuclear IκB function in NF-κB-dependent gene expression, we report an X-ray crystal structure of the IκBζ ankyrin repeat domain in complex with the dimerization domain of the NF-κB p50 homodimer. IκBζ possesses an N-terminal α helix that conveys domain folding stability. Affinity and specificity of the complex depend on a small portion of p50 at the nuclear localization signal. The model suggests that only one p50 subunit supports binding with IκBζ, and biochemical experiments confirm that IκBζ associates with DNA-bound NF-κB p50:RelA heterodimers. Comparisons of IκBζ:p50 and p50:κB DNA complex crystallographic models indicate that structural rearrangement is necessary for ternary complex formation of IκBζ and p50 with DNA.


Assuntos
Modelos Moleculares , Subunidade p50 de NF-kappa B , Ligação Proteica , Multimerização Proteica , Humanos , Sequência de Aminoácidos , Núcleo Celular/metabolismo , Cristalografia por Raios X , DNA/metabolismo , DNA/química , Proteínas I-kappa B/metabolismo , Proteínas I-kappa B/química , Proteínas I-kappa B/genética , Subunidade p50 de NF-kappa B/metabolismo , Subunidade p50 de NF-kappa B/química , Subunidade p50 de NF-kappa B/genética , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelA/química , Fator de Transcrição RelA/genética
2.
Inorg Chem ; 62(28): 10940-10954, 2023 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-37405779

RESUMO

While cancer cells rely heavily upon glycolysis to meet their energetic needs, reducing the importance of mitochondrial oxidative respiration processes, more recent studies have shown that their mitochondria still play an active role in the bioenergetics of metastases. This feature, in combination with the regulatory role of mitochondria in cell death, has made this organelle an attractive anticancer target. Here, we report the synthesis and biological characterization of triarylphosphine-containing bipyridyl ruthenium (Ru(II)) compounds and found distinct differences as a function of the substituents on the bipyridine and phosphine ligands. 4,4'-Dimethylbipyridyl-substituted compound 3 exhibited especially high depolarizing capabilities, and this depolarization was selective for the mitochondrial membrane and occurred within minutes of treatment in cancer cells. The Ru(II) complex 3 exhibited an 8-fold increase in depolarized mitochondrial membranes, as determined by flow cytometry, which compares favorably to the 2-fold increase observed by carbonyl cyanide chlorophenylhydrazone (CCCP), a proton ionophore that shuttles protons across membranes, depositing them into the mitochondrial matrix. Fluorination of the triphenylphosphine ligand provided a scaffold that maintained potency against a range of cancer cells but avoided inducing toxicity in zebrafish embryos at higher concentrations, displaying the potential of these Ru(II) compounds for anticancer applications. This study provides essential information regarding the role of ancillary ligands for the anticancer activity of Ru(II) coordination compounds that induce mitochondrial dysfunction.


Assuntos
Antineoplásicos , Complexos de Coordenação , Rutênio , Animais , 2,2'-Dipiridil , Ligantes , Peixe-Zebra , Mitocôndrias , Rutênio/farmacologia , Rutênio/metabolismo
3.
Inorg Chem ; 60(4): 2178-2187, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-33502194

RESUMO

The specific recognition of AT-rich DNA sequences opens up the door to promising diagnostic and/or therapeutic strategies against gene-related diseases. Here, we demonstrate that amphiphilic PtII complexes of the type [Pt(dmba)(N∧N)]NO3 (dmba = N,N-dimethylbenzylamine-κN, κC; N∧N = dpq (3), dppz (4), and dppn (5)) recognize AT-rich oligonucleotides over other types of DNA, RNA, and model proteins. The crystal structure of 4 shows the presence of significant π-stacking interactions and a distorted coordination sphere of the d8 PtII atom. Complex 5, containing the largest π-conjugated ligand, forms supramolecular assemblies at high concentrations under aqueous environment. However, its aggregation can be promoted in the presence of DNA at concentrations as low as 10 µM in a process that "turns on" its excimer emission around 600 nm. Viscometry, gel electrophoresis, and theoretical calculations demonstrate that 5 binds to minor groove when self-assembled, while the monomers of 3 and 4 intercalate into the DNA. The complexes also inhibit cancer cell growth with low-micromolar IC50 values in 2D tissue culture and suppress tumor growth in 3D tumor spheroids with a multicellular resistance (MCR) index comparable to that of cisplatin.


Assuntos
Complexos de Coordenação/química , DNA/química , Compostos Organoplatínicos/química , Células A549 , Cristalografia por Raios X , Ensaio de Desvio de Mobilidade Eletroforética , Humanos , Substâncias Intercalantes/química , Ligantes , Estrutura Molecular , Análise Espectral/métodos , Estereoisomerismo
4.
Eur J Inorg Chem ; 2021(35): 3611-3621, 2021 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-34539235

RESUMO

The ß-diketone scaffold is a commonly used synthetic intermediate, and is a functional group found in natural products such as curcuminoids. This core structure can also act as a chelating ligand for a variety of metals. In order to assess the potential of this scaffold for medicinal inorganic chemistry, seven different κ2-O,O'-chelating ligands were used to construct Ru(II) complexes with polypyridyl co-ligands, and their biological activity was evaluated. The complexes demonstrated promising structure-dependent cytotoxicity. Three complexes maintained high activity in a tumor spheroid model, and all complexes demonstrated low in vivo toxicity in a zebrafish model. From this series, the best compound exhibited a ~ 30-fold window between cytotoxicity in a 3-D tumor spheroid model and potential in vivo toxicity. These results suggest that κ2-O,O'-ligands can be incorporated into Ru(II)-polypyridyl complexes to create favorable candidates for future drug development.

5.
Inorg Chem ; 59(13): 8882-8892, 2020 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-32530274

RESUMO

Ruthenium(II) complexes developed for photodynamic therapy (PDT) are almost exclusively tris-bidentate systems with C2 or D3 symmetry. This is due to the fact that this structural framework commonly produces long-lived excited states, which, in turn, allow for the generation of large amounts of singlet oxygen (1O2) and other reactive oxygen species. Complexes containing tridentate ligands would be advantageous for biological applications as they are generally achiral (D2d or C2v symmetry), which eliminates the possibility of multiple isomers which could exhibit potentially different interactions with chiral biological entities. However, Ru(II) complexes containing tridentate ligands are rarely studied as candidates for photobiological applications, such as PDT, since they almost exclusively exhibit low quantum yields and very short excited-state lifetimes and, thus, are not capable of generating sufficient 1O2 or engaging in electron transfer reactions. Here, we report a proof-of-concept approach to make bis-tridentate Ru(II) complexes useful for PDT applications by altering their photophysical properties through the inclusion of N-heterocyclic carbene (NHC) ligands. Three NHC and two terpyridine ligands were studied to evaluate the effects of structural and photophysical modulations of bis-substituted Ru(II) complexes. The NHC complexes were found to have superior excited-state lifetimes, 1O2 production, and photocytotoxicity. To the best of our knowledge, these complexes are the most potent light-activated bis-tridentate complexes reported.


Assuntos
Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/efeitos da radiação , Linhagem Celular Tumoral , Complexos de Coordenação/síntese química , Complexos de Coordenação/efeitos da radiação , DNA/metabolismo , Quebras de DNA de Cadeia Simples/efeitos da radiação , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Ligantes , Luz , Estudo de Prova de Conceito , Rutênio/química , Oxigênio Singlete/metabolismo
6.
Biophys J ; 115(7): 1251-1263, 2018 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-30224054

RESUMO

Cytochrome P450BM3 catalyzes the hydroxylation and/or epoxidation of fatty acids, fatty amides, and alcohols. Protein engineering has produced P450BM3 variants capable of accepting drug molecules normally metabolized by human P450 enzymes. The enhanced substrate promiscuity has been attributed to the greater flexibility of the lid of the substrate channel. However, it is not well understood how structurally different and highly polar drug molecules can stably bind in the active site nor how the activity and coupling efficiency of the enzyme may be affected by the lack of enzyme-substrate complementarity. To address these important aspects of non-native small molecule binding, this study investigated the binding of drug molecules with different size, charge, polar surface area, and human P450 affinity on the promiscuous R47L/F87V/L188Q/E267V/F81I pentuple mutant of P450BM3. Binding free energy data and energy decomposition analysis showed that pentuple mutant P450BM3 stably binds (i.e., negative ΔGb°) a broad range of substrate and inhibitor types because dispersion interactions with active site residues overcome unfavorable repulsive and electrostatic effects. Molecular dynamics simulations revealed that 1) acidic substrates tend to disrupt the heme propionate A-K69 salt bridge, which may reduce heme oxidizing ability, and 2) the lack of complementarity leads to high substrate mobility and water density in the active site, which may lead to uncoupling. These factors must be considered in future developments of P450BM3 as a biocatalyst in the large-scale production of drug metabolites.


Assuntos
Proteínas de Bactérias/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Simulação de Dinâmica Molecular , Mutação , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Domínio Catalítico , Sistema Enzimático do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/genética , Heme/metabolismo , Mutagênese Sítio-Dirigida , NADPH-Ferri-Hemoproteína Redutase/química , NADPH-Ferri-Hemoproteína Redutase/genética , Ligação Proteica , Termodinâmica
7.
Mol Pharm ; 15(8): 3404-3416, 2018 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-29865789

RESUMO

Target identification and mechanistic studies of cytotoxic agents are challenging processes that are both time-consuming and costly. Here we describe an approach to mechanism of action studies for potential anticancer compounds by utilizing the simple prokaryotic system, E. coli, and we demonstrate its utility with the characterization of a ruthenium polypyridyl complex [Ru(bpy)2dmbpy2+]. Expression of the photoconvertible fluorescent protein Dendra2 facilitated both high throughput studies and single-cell imaging. This allowed for simultaneous ratiometric analysis of inhibition of protein production and phenotypic investigations. The profile of protein production, filament size and population, and nucleoid morphology revealed important differences between inorganic agents that damage DNA vs more selective inhibitors of transcription and translation. Trace metal analysis demonstrated that DNA is the preferred nucleic acid target of the ruthenium complex, but further studies in human cancer cells revealed altered cell signaling pathways compared to the commonly administrated anticancer agent cisplatin. This study demonstrates E. coli can be used to rapidly distinguish between compounds with disparate mechanisms of action and also for more subtle distinctions within in studies in mammalian cells.


Assuntos
Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Rutênio/farmacologia , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular Tumoral , Cisplatino/farmacologia , Complexos de Coordenação/química , Dano ao DNA/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Ensaios de Triagem em Larga Escala/métodos , Humanos , Biossíntese de Proteínas/efeitos dos fármacos , Rutênio/química , Transcrição Gênica/efeitos dos fármacos
8.
Biochemistry ; 56(1): 240-249, 2017 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-28001373

RESUMO

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) result in the disease cystic fibrosis. Deletion of Phe508, the most prevalent mutation associated with this disease, disrupts trafficking of the protein. Small molecule correctors yield moderate improvements in the trafficking of ΔF508-CFTR to the plasma membrane. It is currently not known if correctors increase the level of trafficking through improved cargo loading of transport vesicles or through direct binding to CFTR. Real-time measurements of trafficking were utilized to identify the mechanistic details of chemical, biochemical, and thermal factors that impact CFTR correction, using the corrector molecule VX-809, a secondary mutation (I539T), and low-temperature conditions. Each individually improved trafficking of ΔF508-CFTR to approximately 10% of wild-type levels. The combination of VX-809 with either low temperature or the I539T mutation increased the amount of CFTR on the plasma membrane to nearly 40%, indicating synergistic activity. The number of vesicles reaching the surface was significantly altered; however, the amount of channel in each vesicle remained the same. Direct binding measurements of VX-809 in native membranes using backscattering interferometry indicate tight binding to CFTR, which occurred in a manner independent of mutation. The similar values obtained for all forms of the channel indicate that the binding site is not compromised or enhanced by these mutations.


Assuntos
Aminopiridinas/metabolismo , Benzodioxóis/metabolismo , Membrana Celular/metabolismo , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Vesículas Transportadoras/metabolismo , Fibrose Cística/genética , Fibrose Cística/metabolismo , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Células HEK293 , Humanos , Concentração de Íons de Hidrogênio , Interferometria/métodos , Microscopia de Fluorescência/métodos , Mutação , Ligação Proteica , Transporte Proteico/genética , Reprodutibilidade dos Testes , Imagem Individual de Molécula/métodos , Temperatura
9.
Inorg Chem ; 56(20): 12214-12223, 2017 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-28949518

RESUMO

Light-activated compounds are powerful tools and potential agents for medical applications, as biological effects can be controlled in space and time. Ruthenium polypyridyl complexes can induce cytotoxic effects through multiple mechanisms, including acting as photosensitizers for singlet oxygen (1O2) production, generating other reactive oxygen species (ROS), releasing biologically active ligands, and creating reactive intermediates that form covalent bonds to biological molecules. A structure-activity relationship (SAR) study was performed on a series of Ru(II) complexes containing isomeric tetramethyl-substituted bipyridyl-type ligands. Three of the ligand systems studied contained strain-inducing methyl groups and created photolabile metal complexes, which can form covalent bonds to biomolecules upon light activation, while the fourth was unstrained and resulted in photostable complexes, which can generate 1O2. The compounds studied included both bis-heteroleptic complexes containing two bipyridine ligands and a third, substituted ligand and tris-homoleptic complexes containing only the substituted ligand. The photophysics, electrochemistry, photochemistry, and photobiology were assessed. Strained heteroleptic complexes were found to be more photoactive and cytotoxic then tris-homoleptic complexes, and bipyridine ligands were superior to bipyrimidine. However, the homoleptic complexes exhibited an enhanced ability to inhibit protein production in live cells. Specific methylation patterns were associated with improved activation with red light, and photolabile complexes were generally more potent cytotoxic agents than the photostable 1O2-generating compounds.


Assuntos
2,2'-Dipiridil/análogos & derivados , 2,2'-Dipiridil/efeitos da radiação , Complexos de Coordenação/efeitos da radiação , Rutênio/química , 2,2'-Dipiridil/síntese química , 2,2'-Dipiridil/farmacologia , Quelantes/química , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Adutos de DNA/efeitos dos fármacos , Quebras de DNA , Replicação do DNA/efeitos dos fármacos , Células HL-60 , Humanos , Ligantes , Luz , Metilação , Biossíntese de Proteínas , Pirimidinas/síntese química , Pirimidinas/química , Pirimidinas/farmacologia , Pirimidinas/efeitos da radiação , Oxigênio Singlete/química , Relação Estrutura-Atividade
10.
Eur J Inorg Chem ; 2017(12): 1687-1694, 2017 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-29200939

RESUMO

Ruthenium complexes capable of light-triggered cytotoxicity are appealing potential prodrugs for photodynamic therapy (PDT) and photoactivated chemotherapy (PACT). Two groups of Ru(II) polypyridyl complexes with 2-(2-pyridyl)-benzazole ligands were synthesized and investigated for their photochemical properties and anticancer activity to compare strained and unstrained systems that are likely to have different biological mechanisms of action. The structure-activity relationship was focused on the benzazole core bioisosterism and replacement of coligands in Ru(II) complexes. Strained compounds rapidly ejected the 2-(2-pyridyl)-benzazole ligand after light irradiation, and possessed strong toxicity in the HL-60 cell line both under dark and light conditions. In contrast, unstrained Ru(II) complexes were non-toxic in the absence of light, induced cytotoxicity at nanomolar concentrations after light irradiation, and are capable of light-induced DNA damage. The 90-220-fold difference in light and dark IC50 values provides a large potential therapeutic window to allow for selective targeting of cells by exposure to light.

11.
Biochemistry ; 55(25): 3594-606, 2016 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-27267136

RESUMO

Cytochrome P450BM3 is a heme-containing enzyme from Bacillus megaterium that exhibits high monooxygenase activity and has a self-sufficient electron transfer system in the full-length enzyme. Its potential synthetic applications drive protein engineering efforts to produce variants capable of oxidizing nonnative substrates such as pharmaceuticals and aromatic pollutants. However, promiscuous P450BM3 mutants often exhibit lower stability, thereby hindering their industrial application. This study demonstrated that the heme domain R47L/F87V/L188Q/E267V/F81I pentuple mutant (PM) is destabilized because of the disruption of hydrophobic contacts and salt bridge interactions. This was directly observed from crystal structures of PM in the presence and absence of ligands (palmitic acid and metyrapone). The instability of the tertiary structure and heme environment of substrate-free PM was confirmed by pulse proteolysis and circular dichroism, respectively. Binding of the inhibitor, metyrapone, significantly stabilized PM, but the presence of the native substrate, palmitic acid, had no effect. On the basis of high-temperature molecular dynamics simulations, the lid domain, ß-sheet 1, and Cys ligand loop (a ß-bulge segment connected to the heme) are the most labile regions and, thus, potential sites for stabilizing mutations. Possible approaches to stabilization include improvement of hydrophobic packing interactions in the lid domain and introduction of new salt bridges into ß-sheet 1 and the heme region. An understanding of the molecular factors behind the loss of stability of P450BM3 variants therefore expedites site-directed mutagenesis studies aimed at developing thermostability.


Assuntos
Bacillus megaterium/enzimologia , Proteínas de Bactérias/química , Sistema Enzimático do Citocromo P-450/química , Metirapona/metabolismo , Proteínas Mutantes/química , Mutação/genética , NADPH-Ferri-Hemoproteína Redutase/química , Ácido Palmítico/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Dicroísmo Circular , Cristalização , Cristalografia por Raios X , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Transporte de Elétrons , Inibidores Enzimáticos/metabolismo , Hidroxilação , Cinética , Modelos Moleculares , Mutagênese Sítio-Dirigida , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , NADPH-Ferri-Hemoproteína Redutase/genética , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Oxirredução , Conformação Proteica
12.
Angew Chem Int Ed Engl ; 54(2): 481-4, 2015 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-25363667

RESUMO

A new approach is presented for the application of single-molecule imaging to membrane receptors through the use of vesicles derived from cells expressing fluorescently labeled receptors. During the isolation of vesicles, receptors remain embedded in the membrane of the resultant vesicles, thus allowing these vesicles to serve as nanocontainers for single-molecule measurements. Cell-derived vesicles maintain the structural integrity of transmembrane receptors by keeping them in their physiological membrane. It was demonstrated that receptors isolated in these vesicles can be studied with solution-based fluorescence correlation spectroscopy (FCS) and can be isolated on a solid substrate for single-molecule studies. This technique was applied to determine the stoichiometry of α3ß4 nicotinic receptors. The method provides the capability to extend single-molecule studies to previously inaccessible classes of receptors.


Assuntos
Proteínas de Membrana/química , Espectrometria de Fluorescência/métodos
13.
Chembiochem ; 15(4): 507-11, 2014 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-24482049

RESUMO

A coupled in vitro transcription and translation (IVTT) assay that uses GFP as a fluorescent reporter allowed the potency of a light-activated cytotoxic ruthenium agent to be quantified. The compound inhibits the function of both DNA and mRNA only upon light activation. The IVTT functional assay provides estimates of potency that are consistent with cellular cytotoxicity values, in marked contrast to the values obtained from traditional DNA-damage assays.


Assuntos
Complexos de Coordenação/metabolismo , DNA/metabolismo , RNA Mensageiro/metabolismo , Rutênio/química , Sistema Livre de Células , Complexos de Coordenação/química , DNA/química , Dano ao DNA , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HeLa , Humanos , Biossíntese de Proteínas , RNA Mensageiro/química , Transcrição Gênica
14.
Inorg Chem ; 53(19): 10030-2, 2014 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-25198057

RESUMO

Two novel strained ruthenium(II) polypyridyl complexes containing a 2,3-dihydro-1,4-dioxino[2,3-f]-1,10-phenanthroline (dop) ligand selectively ejected a methylated ligand when irradiated with >400 nm light. The best compound exhibited a 1880-fold increase in cytotoxicity in human cancer cells upon light-activation and was 19-fold more potent than the well-known chemotherapeutic, cisplatin.


Assuntos
Complexos de Coordenação/química , Compostos Organometálicos/farmacologia , Fenantrolinas/química , Rutênio/química , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células HL-60 , Humanos , Ligantes , Modelos Moleculares , Estrutura Molecular , Compostos Organometálicos/síntese química , Compostos Organometálicos/química , Processos Fotoquímicos , Relação Estrutura-Atividade
15.
Biomolecules ; 14(4)2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38672458

RESUMO

While cytochrome P450 (CYP; P450) enzymes are commonly associated with the metabolism of organic xenobiotics and drugs or the biosynthesis of organic signaling molecules, they are also impacted by a variety of inorganic species. Metallic nanoparticles, clusters, ions, and complexes can alter CYP expression, modify enzyme interactions with reductase partners, and serve as direct inhibitors. This commonly overlooked topic is reviewed here, with an emphasis on understanding the structural and physiochemical basis for these interactions. Intriguingly, while both organometallic and coordination compounds can act as potent CYP inhibitors, there is little evidence for the metabolism of inorganic compounds by CYPs, suggesting a potential alternative approach to evading issues associated with rapid modification and elimination of medically useful compounds.


Assuntos
Inibidores das Enzimas do Citocromo P-450 , Sistema Enzimático do Citocromo P-450 , Sistema Enzimático do Citocromo P-450/metabolismo , Humanos , Inibidores das Enzimas do Citocromo P-450/farmacologia , Inibidores das Enzimas do Citocromo P-450/química , Nanopartículas Metálicas/química , Animais , Metais/química , Metais/metabolismo , Compostos Inorgânicos/química
16.
Biophys J ; 105(4): 975-83, 2013 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-23972849

RESUMO

Deletion of the ß-bulge trigger-loop results in both a switch in the preferred folding route, from the functional loop packing folding route to barrel closure, as well as conversion of the agonist activity of IL-1ß into antagonist activity. Conversely, circular permutations of IL-1ß conserve the functional folding route as well as the agonist activity. These two extremes in the folding-functional interplay beg the question of whether mutations in IL-1ß would result in changes in the populations of heterogeneous folding routes and the signaling activity. A series of topologically equivalent water-mediated ß-strand bridging interactions within the pseudosymmetric ß-trefoil fold of IL-1ß highlight the backbone water interactions that stabilize the secondary and tertiary structure of the protein. Additionally, conserved aromatic residues lining the central cavity appear to be essential for both stability and folding. Here, we probe these protein backbone-water molecule and side chain-side chain interactions and the role they play in the folding mechanism of this geometrically stressed molecule. We used folding simulations with structure-based models, as well as a series of folding kinetic experiments to examine the effects of the F42W core mutation on the folding landscape of IL-1ß. This mutation alters water-mediated backbone interactions essential for maintaining the trefoil fold. Our results clearly indicate that this perturbation in the primary structure alters a structural water interaction and consequently modulates the population of folding routes accessed during folding and signaling activity.


Assuntos
Interleucina-1beta/química , Modelos Moleculares , Dobramento de Proteína , Substituição de Aminoácidos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Mutação , Fenômenos Ópticos , Conformação Proteica , Termodinâmica , Água/química
17.
J Med Chem ; 66(1): 398-412, 2023 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-36520541

RESUMO

Cytochrome P450 1B1 (CYP1B1) is a potential drug target in cancer research that is overexpressed in several solid tumors but is present only at low levels in healthy tissues. Its expression is associated with resistance to common chemotherapeutics, while inhibitors restore efficacy to these drugs in model systems. The majority of CYP1B1 inhibitors are derived from a limited number of scaffolds, and few have achieved outstanding selectivity against other human CYPs, which could impede clinical development. This study explores a new chemical space for CYP1B1 inhibitors using a scaffold-hopping approach and establishes 2,4-diarylthiazoles as a promising framework for further development. From a small library, compound 15 emerged as the lead, with picomolar CYP1B1 inhibition, and over 19,000-fold selectivity against its relative, CYP1A1. To investigate the activity of 15, molecular dynamics, optical spectroscopy, point mutations, and traditional structure-activity relationships were employed and revealed key interactions important for the development of CYP1B1 inhibitors.


Assuntos
Inibidores das Enzimas do Citocromo P-450 , Neoplasias , Humanos , Inibidores das Enzimas do Citocromo P-450/farmacologia , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1B1/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Relação Estrutura-Atividade
18.
J Inorg Biochem ; 238: 112031, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36327501

RESUMO

Photoreactive Ru(II) complexes capable of ejecting ligands have been used extensively for photocaging applications and for the creation of "photocisplatin" reagents. The incorporation of distortion into the structure of the coordination complex lowers the energy of dissociative excited states, increasing the yield of the photosubstitution reaction. While steric clash between ligands induced by adding substituents at the coordinating face of the ligand has been extensively utilized, a lesser known, more subtle approach is to distort the coordination sphere by altering the chelate ring size. Here a systematic study was performed to alter metal-ligand bond lengths, angles, and to cause intraligand distortion by introducing a "linker" atom or group between two pyridine rings. The synthesis, photochemistry, and photobiology of five Ru(II) complexes containing CH2, NH, O, and S-linked dipyridine ligands was investigated. All systems where stable in the dark, and three of the five were photochemically active in buffer. While a clear periodic trend was not observed, this study lays the foundation for the creation of photoactive systems utilizing an alternative type of distortion to facilitate photosubstitution reactions.


Assuntos
Rutênio , Rutênio/química , Ligantes , Fotobiologia , Fotoquímica
19.
RSC Chem Biol ; 4(5): 344-353, 2023 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-37181632

RESUMO

Ruthenium complexes are often investigated as potential replacements for platinum-based chemotherapeutics in hopes of identifying systems with improved tolerability in vivo and reduced susceptibility to cellular resistance mechanisms. Inspired by phenanthriplatin, a non-traditional platinum agent that contains only one labile ligand, monofunctional ruthenium polypyridyl agents have been developed, but until now, few demonstrated promising anticancer activity. Here we introduce a potent new scaffold, based on [Ru(tpy)(dip)Cl]Cl (tpy = 2,2':6',2''-terpyridine and dip = 4,7-diphenyl-1,10-phenanthroline) in pursuit of effective Ru(ii)-based monofunctional agents. Notably, the extension of the terpyridine at the 4' position with an aromatic ring resulted in a molecule that was cytotoxic in several cancer cell lines with sub-micromolar IC50 values, induced ribosome biogenesis stress, and exhibited minimal zebrafish embryo toxicity. This study demonstrates the successful design of a Ru(ii) agent that mimics many of the biological effects and phenotypes seen with phenanthriplatin, despite numerous differences in both the ligands and metal center structure.

20.
J Am Chem Soc ; 134(20): 8324-7, 2012 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-22553960

RESUMO

Strained ruthenium (Ru) complexes have been synthesized and characterized as novel agents for photodynamic therapy (PDT). The complexes are inert until triggered by visible light, which induces ligand loss and covalent modification of DNA. An increase in cytotoxicity of 2 orders of magnitude is observed with light activation in cancer cells, and the compounds display potencies superior to cisplatin against 3D tumor spheroids. The use of intramolecular strain may be applied as a general paradigm to develop light-activated ruthenium complexes for PDT applications.


Assuntos
Antineoplásicos/química , Complexos de Coordenação/química , Fármacos Fotossensibilizantes/química , Rutênio/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/farmacologia , DNA/metabolismo , Humanos , Luz , Neoplasias/tratamento farmacológico , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Rutênio/farmacologia
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