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Biancheri R.,Child Neurology and Psychiatry Unit | Rosano C.,SS Biopolimeri e Proteomica | Denegri L.,Child Neurology and Psychiatry Unit | Lamantea E.,C Besta Neurological Institute | And 4 more authors.
European Journal of Human Genetics | Year: 2013

Homozygous or compound heterozygous mutations in the GJC2 gene, encoding the gap junction protein connexin47 (Cx47), cause the autosomal recessive hypomyelinating Pelizaeus-Merzbacher-like disease (PMLD1, MIM 608804). Although clinical and neuroradiological findings resemble those of the classic Pelizaeus-Merzbacher disease, PMLD patients usually show a greater level of cognitive and motor functions. Unpredictably a homozygous missense GJC2 mutation (p.Glu260Lys) was found in a patient presenting with a very severe clinical picture characterised by congenital nystagmus and severe neurological impairment. Also magnetic resonance imaging was unusually severe, showing an abnormal supra-and infratentorial white matter involvement extending to the spinal cord. The novel p.Glu260Lys (c.778G>A) mutation, occurring in a highly conserved motif (SRPTEK) of the Cx47 extracellular loop-2 domain, was predicted, by modelling analysis, to break a 'salt bridge network', crucial for a proper connexin-connexin interaction to form a connexon, thus hampering the correct formation of the connexon pore. The same structural analysis, extended to the previously reported missense mutations, predicted that most changes were expected to have less severe impact on protein functions, correlating with the mild PMLD1 form of the patients. Our study expands the spectrum of PMLD1 and provides evidence that the extremely severe clinical and neuroradiological PMLD1 form of our patient likely correlates with the predicted impairment of gap junction channel assembly resulting from the detrimental effect of the new p.Glu260Lys mutant allele on Cx47 protein. © 2013 Macmillan Publishers Limited. All rights reserved.

Mangerini R.,Science Oncologia Medica B | Romano P.,Science Anatomia e Citoistologia Patologica | Facchiano A.,CNR Institute of Food Sciences | Damonte G.,University of Genoa | And 5 more authors.
Analytical Biochemistry | Year: 2011

Although most time-of-flight (TOF) mass spectrometers come equipped with vacuum matrix-assisted laser desorption/ionization (MALDI) sources, the atmospheric pressure MALDI (API-MALDI) source is an attractive option because of its ability to be coupled to a wide range of analyzers. This article describes the use of an API-MALDI source coupled to a TOF mass spectrometer for evaluation of the effects of medium- and long-term storage on peptidomic profiles of cryopreserved serum samples from healthy women. Peptides were purified using superparamagnetic beads either from fresh sera or after serum storage at -80 °C for 18 months or at -20 °C for 8 years. Data were preprocessed using newly developed bioinformatic tools and then were subjected to statistical analysis and class prediction. The analyses showed a dramatic effect of storage on the abundance of several peptides such as fibrinopeptides A and B, complement fractions, bradykinin, and clusterin, indicated by other authors as disease biomarkers. Most of these results were confirmed by shadow clustering analysis, able to classify each sample in the correct group. In addition to demonstrating the suitability of the API-MALDI technique for peptidome profiling studies, our data are of relevance for retrospective studies that involve frozen sera stored for many years in biobanks. © 2011 Elsevier Inc.

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