Insights into Protein Aggregation by NMR Characterization of Insoluble SH3 Mutants Solubilized in Salt-Free Water

Protein aggregation in vivo has been extensively associated with a large spectrum of human diseases. On the other hand, mechanistic insights into protein aggregation in vitro were incomplete due to the inability in solubilizing insoluble proteins for high-resolution biophysical investigations. However, a new avenue may be opened up by our recent discovery that previously-thought insoluble proteins can in fact be solubilized in salt-free water. Here we use this approach to study the NMR structural and dynamic properties of an insoluble SH3 mutant with a naturally-occurring insertion of Val22 at the tip of the diverging turn. The obtained results reveal: 1) regardless of whether the residue is Val, Ala, Asp or Arg, the insertion will render the first hNck2 SH3 domain to be insoluble in buffers. Nevertheless, all four mutants could be solubilized in salt-free water and appear to be largely unfolded as evident from their CD and NMR HSQC spectra. 2) Comparison of the chemical shift deviations reveals that while in V22-SH3 the second helical region is similarly populated as in the wild-type SH3 at pH 2.0, the first helical region is largely unformed. 3) In V22-SH3, many non-native medium-range NOEs manifest to define non-native helical conformations. In the meanwhile a small group of native-like long-range NOEs still persists, indicating the existence of a rudimentary native-like tertiary topology. 4) Although overall, V22-SH3 has significantly increased backbone motions on the ps-ns time scale, some regions still own restricted backbone motions as revealed by analyzing 15N relaxation data. Our study not only leads to the establishment of the first high-resolution structural and dynamic picture for an insoluble protein, but also shed more light on the molecular events for the nonhierarchical folding mechanism. Furthermore, a general mechanism is also proposed for in vivo protein aggregation triggered by the genetic mutation and posttranslational modification

Alimentos na consistência líquida e deglutição: uma revisão crítica da literatura Food in liquid consistency and deglutition: a critical review of the literature

O objetivo da presente revisão de literatura foi analisar artigos científicos internacionais publicados sobre a fisiologia da deglutição de alimentos líquidos nas fases oral e faríngea. A metodologia empregada envolveu a formulação da pergunta; localização e seleção dos estudos; avaliação crítica dos artigos; conforme os preceitos do Cochrane Handbook. Foram identificados 185 artigos, dos quais se excluiu 141 por não relacionarem-se diretamente ao tema e analisou-se 29 estudos. As pesquisas estão fortemente relacionadas às formas de identificação de disfagia e não as características proporcionadas pela deglutição de diferentes consistências. Quanto à metodologia empregada nos artigos analisados observa-se que na maioria dos estudos não há grupo-controle. Os grupos estudados são heterogêneos, principalmente quando considerando indivíduos com alterações neurológicas, além disso, não há pareamento de idade na maioria dos estudos. Dessa forma, os achados desta revisão demonstram que há dificuldade na aplicabilidade clínica dos achados científicos, dificultando a prática baseada evidências.<br>The purpose of the present literature review was to analyze international scientific papers published on the physiology of swallowing fluids in the oral and pharyngeal phases. The employed methodology involved the formulation of the question, location and selection of studies, critical assessment of the manuscripts, according to the concepts of the Cochrane Handbook. We identified 185 articles, out of which 141 were excluded for not being directly related to the theme. Twenty-nine studies were analyzed. The researchers are strongly related to ways of identifying dysphagia and not the features offered by swallowing various consistencies. Regarding the methodology employed in the reviewed articles it is observed that in most studies there are no control groups. The studied groups are heterogeneous, especially when considering individuals with neurological disorders. In addition, subjects are not paired by age. Thus, the findings of this review indicate that there is great difficulty for clinicians to apply in their daily practice scientific findings, which in turn makes evidence-based practice very limited