RESUMEN
Chiral control of crystallization has ample precedent in the small-molecule world, but relatively little is known about the role of chirality in protein crystallization. In this study, lysozyme was crystallized in the presence of the chiral additive 2-methyl-2,4-pentanediol (MPD) separately using the R and S enantiomers as well as with a racemic RS mixture. Crystals grown with (R)-MPD had the most order and produced the highest resolution protein structures. This result is consistent with the observation that in the crystals grown with (R)-MPD and (RS)-MPD the crystal contacts are made by (R)-MPD, demonstrating that there is preferential interaction between lysozyme and this enantiomer. These findings suggest that chiral interactions are important in protein crystallization.
Asunto(s)
Glicoles/química , Muramidasa/química , Cristalografía por Rayos X , Estructura Terciaria de ProteínaRESUMEN
The endocannabinoid CB2 receptor system has been implicated in the neuropathology of Alzheimer's disease (AD). In order to investigate the impact of the CB2 receptor system on AD pathology, a colony of mice with a deleted CB2 receptor gene, CNR2, was established on a transgenic human mutant APP background for pathological comparison with CB2 receptor-sufficient transgenic mice. J20 APP (PDGFB-APPSwInd) mice were bred over two generations with CNR2(-/-) (Cnr2(tm1Dgen)/J) mice to produce a colony of J20 CNR2(+/+) and J20 CNR2(-/-) mice. Seventeen J20 CNR2(+/+) mice (12 females, 5 males) and 16 J20 CNR2(-/-) mice (11 females, 5 males) were killed at 12 months, and their brains were interrogated for AD-related pathology with both biochemistry and immunocytochemistry (ICC). In addition to amyloid-dependent endpoints such as soluble Aß production and plaque deposition quantified with 6E10 staining, the effect of CB2 receptor deletion on total soluble mouse tau production was assayed by using a recently developed high-sensitivity assay. Results revealed that soluble Aß42 and plaque deposition were significantly increased in J20 CNR2(-/-) mice relative to CNR2(+/+) mice. Microgliosis, quantified with ionized calcium-binding adapter molecule 1 (Iba-1) staining, did not differ between groups, whereas plaque associated microglia was more abundant in J20 CNR2(-/-) mice. Total tau was significantly suppressed in J20 CNR2(-/-) mice relative to J20 CNR2(+/+) mice. The results confirm the constitutive role of the CB2 receptor system both in reducing amyloid plaque pathology in AD and also support tehpotential of cannabinoid therapies targeting CB2 to reduce Aß; however, the results suggest that interventions may have a divergent effect on tau pathology.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Receptor Cannabinoide CB2/metabolismo , Proteínas tau/metabolismo , Enfermedad de Alzheimer/patología , Precursor de Proteína beta-Amiloide/genética , Animales , Encéfalo/metabolismo , Encéfalo/patología , Proteínas de Unión al Calcio/metabolismo , Agonistas de Receptores de Cannabinoides/farmacología , Cannabinoides/farmacología , Proteínas de Unión al ADN , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones Transgénicos , Proteínas de Microfilamentos/metabolismo , Microglía/metabolismo , Microglía/patología , Placa Amiloide/metabolismo , Placa Amiloide/patología , Proteínas del Grupo Polycomb , Receptor Cannabinoide CB2/agonistas , Receptor Cannabinoide CB2/genética , Factores de Transcripción/metabolismoRESUMEN
The endocannabinoid CB2 receptor system has been implicated in the neuropathology of Alzheimer's disease (AD). In order to investigate the impact of the CB2 receptor system on AD pathology, a colony of mice with a deleted CB2 receptor gene, CNR2, was established on a transgenic human mutant APP background for pathological comparison with CB2 receptor-sufficient transgenic mice. J20 APP (PDGFB-APPSwInd) mice were bred over two generations with CNR2â»/â» (Cnr2(tm1Dgen)/J) mice to produce a colony of J20 CNR2âº/⺠and J20 CNR2â»/â» mice. Seventeen J20 CNR2âº/⺠mice (12 females, 5 males) and 16 J20 CNR2â»/â» mice (11 females, 5 males) were killed at 12 months, and their brains were interrogated for AD-related pathology with both biochemistry and immunocytochemistry (ICC). In addition to amyloid-dependent endpoints such as soluble Aß production and plaque deposition quantified with 6E10 staining, the effect of CB2 receptor deletion on total soluble mouse tau production was assayed by using a recently developed high-sensitivity assay. Results revealed that soluble Aß42 and plaque deposition were significantly increased in J20 CNR2â»/â» mice relative to CNR2âº/⺠mice. Microgliosis, quantified with ionized calcium-binding adapter molecule 1 (Iba-1) staining, did not differ between groups, whereas plaque associated microglia was more abundant in J20 CNR2â»/â» mice. Total tau was significantly suppressed in J20 CNR2â»/â» mice relative to J20 CNR2âº/⺠mice. The results confirm the constitutive role of the CB2 receptor system both in reducing amyloid plaque pathology in AD and also support tehpotential of cannabinoid therapies targeting CB2 to reduce Aß; however, the results suggest that interventions may have a divergent effect on tau pathology.