RESUMEN
Little is known about the healthy aging of the circadian timing system under natural living conditions. This study explores changes in the circadian rhythm of distal skin temperature (DST) with aging and relates these changes to sleep-wake timing and environmental influences. DST, sleep-wake timing, 24-h light exposure, and physical activity were measured and averaged over seven consecutive days using temperature sensors, actigraphy with a light meter, and sleep diaries in 35 healthy older women (60-79 years) and 30 young women (20-34 years). Circadian rhythm characteristics, describing strength (amplitude) and timing (acrophase) of the DST rhythm, were calculated using cosinor analysis. The older adults displayed an 18-19% smaller amplitude and a 66-73 min earlier acrophase (peak time) for DST rhythm than the young adults, indicating a weaker and phase-advanced DST rhythm. The phase advance for DST was not due to an earlier evening increase, but to a shorter nocturnal plateau period. Daytime light exposure inversely affected strength (amplitude) but not phasing of the DST rhythm in older adults. The DST rhythm was 3.5 times more advanced than the sleep-wake rhythm, showing an altered phase relationship (phase angle) between both rhythms with aging. The phase angle was more heterogeneous among older adults, showing differential aging. The phase advance for DST rhythm and the altered and heterogeneous phase relationship between DST and sleep-wake rhythms were not related to ambient light exposure and the physical activity of older adults. This suggests that healthy aging of the circadian system might be due to endogenous mechanisms such as an internal rearrangement rather than external influences.
RESUMEN
The formation of oligomers of the amyloid-ß peptide plays a key role in the onset of Alzheimer's disease. We describe herein the investigation of disease-relevant small amyloid-ß oligomers by mass spectrometry and ion mobility spectrometry, revealing functionally relevant structural attributes. In particular, we can show that amyloid-ß oligomers develop in two distinct arrangements leading to either neurotoxic oligomers and fibrils or non-toxic amorphous aggregates. Comprehending the key-attributes responsible for those pathways on a molecular level is a pre-requisite to specifically target the peptide's tertiary structure with the aim to promote the emergence of non-toxic aggregates. Here, we show for two fibril inhibiting ligands, an ionic molecular tweezer and a hydrophobic peptide that despite their different interaction mechanisms, the suppression of the fibril pathway can be deduced from the disappearance of the corresponding structure of the first amyloid-ß oligomers.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/química , Péptidos beta-Amiloides/metabolismo , Enfermedad de Alzheimer/genética , Péptidos beta-Amiloides/genética , Humanos , Ligandos , Espectrometría de Masas , Modelos Moleculares , Agregado de ProteínasRESUMEN
The peptide sequence KLVFF resembles the hydrophobic core of the Aß peptide known to form amyloid plaques in Alzheimer's disease. Starting from its retro-inverso peptide, we have synthesized three generations of peptidomimetics. Step by step natural amino acids have been replaced by aromatic building blocks accessible from the Pd-catalyzed Catellani reaction. The final compound 18 is stable against proteolytic decay and largely prevents the aggregation of Aß1-42 over extended periods of time. The activity of the new inhibitors was tested first by fluorescence correlation spectroscopy. For closer examination of compound 18, additional techniques were also applied: laser-induced liquid bead ion desorption mass spectrometry, confocal laser scanning microscopy, thioflavin T fluorescence, and gel electrophoresis. Compound 18 not only retards the aggregation of chemically synthesized Aß but also can partially dissolve the oligomeric structures. Thioflavin binding mature fibrils, however, seem to resist the inhibitor.
Asunto(s)
Péptidos beta-Amiloides , Fragmentos de Péptidos , Peptidomiméticos/química , Peptidomiméticos/farmacología , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Humanos , Lactato Deshidrogenasas/genética , Lactato Deshidrogenasas/metabolismo , Estructura Molecular , Fragmentos de Péptidos/química , Unión ProteicaRESUMEN
The vibrational fingerprints of hydrogen-bonding associated with the adenine-thymine (A-T) Watson-Crick (WC) base pair have been identified in an infrared study of the A-T mimics 4-aminopyrimidine-1-methylthymine (4APM-1MT) and 4-aminopyrimidine-6-methyl-4-pyrimidinone (4APM-M4PMN) in the gas-phase. The IR vibrational spectra were measured via a double resonance scheme utilizing femtosecond multiphoton ionization. The changes in the molecular structure, anharmonic vibrational parameters, and the assignment of the observed vibrational spectra in the NH/CH stretch region were investigated by carrying out high-level theoretical calculations of the anharmonic spectra. The experimental observations and theoretical calculations indicate that the hydrogen bonds associated with WC base-pairing are relatively stronger than those associated with reverse WC (rWC) base pairing. This is manifested in a more pronounced red-shift of the H-bonded vibrational modes associated with the WC as compared with the rWC base-pairing. An analysis of the factors contributing to the anharmonicity of the vibrational modes associated with H-bonding reveals that the magnitude of the off-diagonal anharmonic coupling of the H-bonded -NH2 stretch and the -NH2 bend is much smaller in WC base-pairing than in the corresponding rWC base-pairing. The chemical and biological implications of these results, especially in the context of using vibrational spectroscopy as a tool for identifying the signatures of nucleotide base vibrations is addressed.
Asunto(s)
Pirimidinas/química , Pirimidinonas/química , Timina/análogos & derivados , Emparejamiento Base , Timina/química , VibraciónRESUMEN
In the present work, the recently developed laser-induced liquid bead ion desorption mass spectrometry (LILBID MS) is applied as a novel technique to study Aß oligomerization, thought to be crucial in Alzheimer's disease (AD). The characterization of the earliest nucleation events of this peptide necessitates the application of several techniques to bridge the gap between small oligomers and large fibrils. We precisely monitored in time the transformation of monomeric Aß (1-42) into oligomeric Aß(n) (n < 20) and its dependence on concentration and agitation. The distribution shows signs of the hexamer being crucial in the assembly process. The intensity of the monomer decreases in time with a time constant of about 9 h. After a lag time of around 10 h, a phase transition occurred in which the total ion current of the oligomers goes to nearly zero. In this late stage of aggregation, protofibrils are formed and mass spectrometry is no longer sensitive. Here fluorescence correlation spectroscopy (FCS) and transmission electron microscopy (TEM) are complementary tools for detection and size characterization of these large species. We also utilized the oligomers of Aß (1-42) as a model of the corresponding in vivo process to screen the efficacy and specificity of small molecule inhibitors of oligomerization. The LILBID results are in excellent agreement with condensed phase behavior determined in other studies. Our data identified LILBID MS as a powerful technique that will advance the understanding of peptide oligomerization in neurodegenerative diseases and represents a powerful tool for the identification of small oligomerization inhibitors.
Asunto(s)
Péptidos beta-Amiloides/química , Rayos Láser , Espectrometría de Masas/métodos , Fragmentos de Péptidos/química , Multimerización de Proteína , Secuencia de Aminoácidos , Benzotiazoles , Humanos , Concentración de Iones de Hidrógeno , Cinética , Datos de Secuencia Molecular , Multimerización de Proteína/efectos de los fármacos , Estructura Secundaria de Proteína , Bibliotecas de Moléculas Pequeñas/farmacología , Espectrometría de Fluorescencia , Tiazoles/químicaRESUMEN
We report the infrared spectrum of the 4-aminobenzimidazole-1-methylthymine (4ABI:1MT) heterodimer, detected by femtosecond multiphoton ionization. Based on calculations of both the harmonic and the anharmonic frequencies, the observed vibrational spectrum is assigned to a structure that mimics the Hoogsteen base pairing of adenine and thymine. A notable observation made in the course of this study is that there is a significant imbalance in the observed strengths of the H-bonds. While the N···H-N bond reveals a large red shift of >700 cm(-1) for the NH stretch frequency, the N-H···O bond is characterized by only a 50 cm(-1) shift. The importance of this observation in the formation of Hoogsteen duplexes by thymine-based oligonucleotides is discussed.