Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 4 de 4
Filtrar
Más filtros

Banco de datos
Tipo del documento
Asunto de la revista
Intervalo de año de publicación
1.
Proc Natl Acad Sci U S A ; 120(49): e2203241120, 2023 12 05.
Artículo en Inglés | MEDLINE | ID: mdl-38015839

RESUMEN

The Lysinibacillus sphaericus proteins Tpp49Aa1 and Cry48Aa1 can together act as a toxin toward the mosquito Culex quinquefasciatus and have potential use in biocontrol. Given that proteins with sequence homology to the individual proteins can have activity alone against other insect species, the structure of Tpp49Aa1 was solved in order to understand this protein more fully and inform the design of improved biopesticides. Tpp49Aa1 is naturally expressed as a crystalline inclusion within the host bacterium, and MHz serial femtosecond crystallography using the novel nanofocus option at an X-ray free electron laser allowed rapid and high-quality data collection to determine the structure of Tpp49Aa1 at 1.62 Å resolution. This revealed the packing of Tpp49Aa1 within these natural nanocrystals as a homodimer with a large intermolecular interface. Complementary experiments conducted at varied pH also enabled investigation of the early structural events leading up to the dissolution of natural Tpp49Aa1 crystals-a crucial step in its mechanism of action. To better understand the cooperation between the two proteins, assays were performed on a range of different mosquito cell lines using both individual proteins and mixtures of the two. Finally, bioassays demonstrated Tpp49Aa1/Cry48Aa1 susceptibility of Anopheles stephensi, Aedes albopictus, and Culex tarsalis larvae-substantially increasing the potential use of this binary toxin in mosquito control.


Asunto(s)
Bacillaceae , Bacillus , Culex , Plaguicidas , Animales , Bacillaceae/química , Bacillaceae/metabolismo , Control de Mosquitos , Larva/metabolismo
2.
Nat Commun ; 15(1): 3827, 2024 May 07.
Artículo en Inglés | MEDLINE | ID: mdl-38714735

RESUMEN

The main protease (Mpro) of SARS-CoV-2 is critical for viral function and a key drug target. Mpro is only active when reduced; turnover ceases upon oxidation but is restored by re-reduction. This suggests the system has evolved to survive periods in an oxidative environment, but the mechanism of this protection has not been confirmed. Here, we report a crystal structure of oxidized Mpro showing a disulfide bond between the active site cysteine, C145, and a distal cysteine, C117. Previous work proposed this disulfide provides the mechanism of protection from irreversible oxidation. Mpro forms an obligate homodimer, and the C117-C145 structure shows disruption of interactions bridging the dimer interface, implying a correlation between oxidation and dimerization. We confirm dimer stability is weakened in solution upon oxidation. Finally, we observe the protein's crystallization behavior is linked to its redox state. Oxidized Mpro spontaneously forms a distinct, more loosely packed lattice. Seeding with crystals of this lattice yields a structure with an oxidation pattern incorporating one cysteine-lysine-cysteine (SONOS) and two lysine-cysteine (NOS) bridges. These structures further our understanding of the oxidative regulation of Mpro and the crystallization conditions necessary to study this structurally.


Asunto(s)
Dominio Catalítico , Proteasas 3C de Coronavirus , Cisteína , Disulfuros , Oxidación-Reducción , SARS-CoV-2 , Disulfuros/química , Disulfuros/metabolismo , SARS-CoV-2/metabolismo , SARS-CoV-2/química , Proteasas 3C de Coronavirus/metabolismo , Proteasas 3C de Coronavirus/química , Cisteína/química , Cisteína/metabolismo , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Multimerización de Proteína , COVID-19/virología
3.
Commun Biol ; 6(1): 1058, 2023 10 18.
Artículo en Inglés | MEDLINE | ID: mdl-37853179

RESUMEN

Several drug screening campaigns identified Calpeptin as a drug candidate against SARS-CoV-2. Initially reported to target the viral main protease (Mpro), its moderate activity in Mpro inhibition assays hints at a second target. Indeed, we show that Calpeptin is an extremely potent cysteine cathepsin inhibitor, a finding additionally supported by X-ray crystallography. Cell infection assays proved Calpeptin's efficacy against SARS-CoV-2. Treatment of SARS-CoV-2-infected Golden Syrian hamsters with sulfonated Calpeptin at a dose of 1 mg/kg body weight reduces the viral load in the trachea. Despite a higher risk of side effects, an intrinsic advantage in targeting host proteins is their mutational stability in contrast to highly mutable viral targets. Here we show that the inhibition of cathepsins, a protein family of the host organism, by calpeptin is a promising approach for the treatment of SARS-CoV-2 and potentially other viral infections.


Asunto(s)
COVID-19 , Humanos , SARS-CoV-2/metabolismo , Catepsinas , Antivirales/farmacología , Antivirales/uso terapéutico , Antivirales/química , Inhibidores de Proteasas/farmacología , Cisteína Endopeptidasas/metabolismo
4.
Science ; 382(6674): 1015-1020, 2023 12.
Artículo en Inglés | MEDLINE | ID: mdl-38033070

RESUMEN

Photolyase is an enzyme that uses light to catalyze DNA repair. To capture the reaction intermediates involved in the enzyme's catalytic cycle, we conducted a time-resolved crystallography experiment. We found that photolyase traps the excited state of the active cofactor, flavin adenine dinucleotide (FAD), in a highly bent geometry. This excited state performs electron transfer to damaged DNA, inducing repair. We show that the repair reaction, which involves the lysis of two covalent bonds, occurs through a single-bond intermediate. The transformation of the substrate into product crowds the active site and disrupts hydrogen bonds with the enzyme, resulting in stepwise product release, with the 3' thymine ejected first, followed by the 5' base.


Asunto(s)
Desoxirribodipirimidina Fotoliasa , Cristalografía , Desoxirribodipirimidina Fotoliasa/química , Desoxirribodipirimidina Fotoliasa/metabolismo , Reparación del ADN , Daño del ADN , Transporte de Electrón
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA