Protein instability and functional defects caused by mutations of dihydro-orotate dehydrogenase in Miller syndrome patients

Synopsis Miller syndrome is a recessive inherited disorder characterized by postaxial acrofacial dysostosis. It is caused by dysfunction of the DHODH (dihydroorotate dehydrogenase) gene, which encodes a key enzyme in the pyrimidine de novo biosynthesis pathway and is localized at mitochondria intermembrane space. We investigated the consequence of three missense mutations, G202A, R346W and R135C of DHODH, which were previously identified in patients with Miller syndrome. First, we established HeLa cell lines stably expressing DHODH with Miller syndrome-causative mutations: G202A, R346W and R135C. These three mutant proteins retained the proper mitochondrial localization based on immunohistochemistry and mitochondrial subfractionation studies. The G202A, R346W DHODH proteins showed reduced protein stability. On the other hand, the third one R135C, in which the mutation lies at the ubiquinone-binding site, was stable but possessed no enzymatic activity. In conclusion, the G202A and R346W mutation causes deficient protein stability, and the R135C mutation does not affect stability but impairs the substrate-induced enzymatic activity, suggesting that impairment of DHODH activity is linked to the Miller syndrome phenotype

Purification, crystallization and preliminary X-ray analysis of a deletion mutant of a major buckwheat allergen

A 16 kDa buckwheat protein (BWp16) is a major allergen responsible for immediate hypersensitivity reactions including anaphylaxis. An immunologically active mutant of BWp16 was prepared and a three-wavelength MAD data set was collected from a crystal of selenomethionine-labelled mutant protein

Association of Tannins and Related Polyphenols with the Cyclic Peptide Gramicidin S

The association of 10 different tannins and related polyphenols with gramicidin S, a cyclic peptide having a rigid β-turn structure, has been examined using 1H-NMR spectroscopy. In the presence of pentagalloylglucose and epigallocatechin-3-O-gallate, the proton signals due to proline and the adjacent phenylalanine moieties selectively shifted to up field, suggesting a regioselective association with the β-turn structure. The association was also supported by the observation of intermolecular nuclear Overhauser effects between epigallocatechin-3-O-gallate and the peptide. In contrast, ellagitannins, biogenetically derived from pentagalloylglucose, showed small and non-selective chemical shift changes, suggesting that interaction with these tannins is relatively weak. The hydrophobicity of the tannin molecules and the steric hindrance of the interaction site are thought to be important in the association

Households with Insufficient Bednets in a Village with Sufficient Bednets: Evaluation of Household Bednet Coverage Using Bednet Distribution Index in Xepon District, Lao PDR

In Lao PDR, the National Malaria Control Program (NMCP) evaluates bednet coverage, often at the village level, using a coverage target of one net per 2.5 (or fewer) persons in a given population. However, in villages that meet the target, not all households necessarily meet the target or utilize all available bednets. This study explored households that fell short of the target and household utilization of bednets in villages that met the target of bednet coverage set by the NMCP. The person per net ratio (PPNR), which is defined as the population divided by the number of available bednets in a household/village, was used to determine whether a household/ village met the NMCP target. Using a household survey, we collected and analyzed the data of 635 households in 17 villages in Xepon district in 2012. Households that fell short of the target (households with a PPNR of > 2.5 or no bednet) existed in every village. The proportion of these households differed greatly among the villages, ranging from 3.4?50%, with some households falling far short. Of the 635 households, 275 (43.5%) had at least one bednet that was not being used on the night preceding the survey and 131 (20.6%) had at least two. In conclusion, in villages that met the NMCP target, a considerable number of households fell short of the target, and the available bednets were not fully utilized in many of the surveyed households

Tbx1, a gene encoded in 22q11.2 copy number variant, is a link between alterations in fimbria myelination and cognitive speed in mice.

Copy number variants (CNVs) have provided a reliable entry point to identify the structural correlates of atypical cognitive development. Hemizygous deletion of human chromosome 22q11.2 is associated with impaired cognitive function; however, the mechanisms by which the CNVs contribute to cognitive deficits via diverse structural alterations in the brain remain unclear. This study aimed to determine the cellular basis of the link between alterations in brain structure and cognitive functions in mice with a heterozygous deletion of Tbx1, one of the 22q11.2-encoded genes. Ex vivo whole-brain diffusion-tensor imaging (DTI)-magnetic resonance imaging (MRI) in Tbx1 heterozygous mice indicated that the fimbria was the only region with significant myelin alteration. Electron microscopic and histological analyses showed that Tbx1 heterozygous mice exhibited an apparent absence of large myelinated axons and thicker myelin in medium axons in the fimbria, resulting in an overall decrease in myelin. The fimbria of Tbx1 heterozygous mice showed reduced mRNA levels of Ng2, a gene required to produce oligodendrocyte precursor cells. Moreover, postnatal progenitor cells derived from the subventricular zone, a source of oligodendrocytes in the fimbria, produced fewer oligodendrocytes in vitro. Behavioral analyses of these mice showed selectively slower acquisition of spatial memory and cognitive flexibility with no effects on their accuracy or sensory or motor capacities. Our findings provide a genetic and cellular basis for the compromised cognitive speed in patients with 22q11.2 hemizygous deletion