Macrocytosis in Mitochondrial DNA Deletion Syndromes.

Large single mitochondrial DNA (mtDNA) deletion syndrome is a rare inborn error of metabolism with variable heteroplasmy levels and clinical phenotype among affected individuals. Chronic progressive external ophthalmoplegia (CPEO) is the most common phenotype in adults with this form of mitochondrial disease [J Intern Med. 2020;287(6):592-608 and Biomed Rep.

A simple method modification to increase separation of 2- and 3-hydroxyglutaric acid by GC-MS for clinical urine organic acids analysis.

Urine organic acids profiling by gas chromatography-mass spectrometry (GC-MS) is routinely performed in hospital biochemical genetics laboratories for the investigation of inborn errors of metabolism. In particular, accurate identification of urinary levels of 3-hydroxyglutaric acid (3-OHGA) is important for diagnosing glutaric aciduria type 1 (GA1), but can be challenging by routine GC-MS profiling analysis due to co-elution and spectral similarity with the isomer 2-hydroxyglutaric acid (2-OHGA). To improve analytical specificity, unique ions were selected and a simple second-tier reinjection method was developed to enhance the chromatographic separation of the 2- and 3-OHGA isomers and potential unknown interferences.

Analysis of 2-methylcitric acid, methylmalonic acid, and total homocysteine in dried blood spots by LC-MS/MS for application in the newborn screening laboratory: A dual derivatization approach.

Inborn errors of propionate, cobalamin and methionine metabolism are targets for Newborn Screening (NBS) in most programs world-wide, and are primarily screened by analyzing for propionyl carnitine (C3) and methionine in dried blood spot (DBS) cards using tandem mass spectrometry (MS/MS). Single-tier NBS approaches using C3 and methionine alone lack specificity, which can lead to an increased false-positive rate if conservative cut-offs are applied to minimize the risk of missing cases. Implementation of liquid chromatography tandem mass spectrometry (LC-MS/MS) second-tier testing for 2-methylcitric acid (MCA), methylmalonic acid (MMA), and homocysteine (HCY) from the same DBS card can improve disease screening performance by reducing the false-positive rate and eliminating the need for repeat specimen collection.

Molecular Diagnosis of Pompe Disease in the Genomic Era: Correlation with Acid Alpha-Glucosidase Activity in Dried Blood Spots.

Measurement of alpha-glucosidase activity on dried blood spots has been the main method to screen for Pompe disease, but a paradigm shift has been observed in recent years with the incorporation of gene panels and exome sequencing in molecular diagnostic laboratories. An 89-gene panel has been available to Canadian physicians since 2017 and was analyzed in 2030 patients with a suspected muscle disease. Acid alpha-glucosidase activity was measured in parallel in dried blood spots from 1430 patients.

Congenital lactic acidosis, cerebral cysts and pulmonary hypertension in an infant with FOXRED1 related complex 1 deficiency.

Mitochondrial complex I is encoded by 38 nuclear-encoded and 7 mitochondrial-encoded genes. is one of the 13 additional nuclear genes known as assembly factors. So far, four patients have been described with complex I deficiency caused by autosomal recessive mutations in Here, we report the fifth patient with related complex 1 deficiency presenting with prenatal onset of bilateral periventricular cysts, congenital lactic acidosis, and persistent life-limiting pulmonary hypertension.

Gene Expression Profiling of Endoplasmic Reticulum Stress in Hepatitis C Virus-Containing Cells Treated with an Inhibitor of Protein Disulfide Isomerases.

Protein disulfide isomerases (PDIs) catalyze disulfide bond formation between protein cysteine residues during protein folding in the endoplasmic reticulum (ER) lumen and are essential for maintaining ER homoeostasis. The life cycle of the hepatitis C virus (HCV) is closely associated with the ER. Synthesis and maturation of HCV proteins occur in the ER membrane and are mediated by multiple host cell factors that include also PDI.

Congenital lactic acidosis, cerebral cysts and pulmonary hypertension in an infant with FOXRED1 related complex I deficiency.

Mitochondrial complex I is encoded by 38 nuclear-encoded and 7 mitochondrial-encoded genes. is one of the 13 additional nuclear genes known as assembly factors. So far, four patients have been described with complex I deficiency caused by autosomal recessive mutations in Here, we report the fifth patient with related complex 1 deficiency presenting with prenatal onset of bilateral periventricular cysts, congenital lactic acidosis, and persistent life-limiting pulmonary hypertension.

Sialic acid catabolism by N-acetylneuraminate pyruvate lyase is essential for muscle function.

Sialic acids are important components of glycoproteins and glycolipids essential for cellular communication, infection, and metastasis. The importance of sialic acid biosynthesis in human physiology is well illustrated by the severe metabolic disorders in this pathway. However, the biological role of sialic acid catabolism in humans remains unclear.

Consensus recommendations for the diagnosis, treatment and follow-up of inherited methylation disorders.

Inherited methylation disorders are a group of rarely reported, probably largely underdiagnosed disorders affecting transmethylation processes in the metabolic pathway between methionine and homocysteine. These are methionine adenosyltransferase I/III, glycine N-methyltransferase, S-adenosylhomocysteine hydrolase and adenosine kinase deficiencies. This paper provides the first consensus recommendations for the diagnosis and management of methylation disorders.

Exome Sequencing and the Management of Neurometabolic Disorders.

Background: Whole-exome sequencing has transformed gene discovery and diagnosis in rare diseases. Translation into disease-modifying treatments is challenging, particularly for intellectual developmental disorder. However, the exception is inborn errors of metabolism, since many of these disorders are responsive to therapy that targets pathophysiological features at the molecular or cellular level.

Structure-based mutagenic analysis of mechanism and substrate specificity in mammalian glycosyltransferases: porcine ST3Gal-I.

Sialyltransferases (STs) play essential roles in signaling and in the cellular recognition processes of mammalian cells by selectively installing cell-surface sialic acids in an appropriate manner both temporally and organ-specifically. The availability of the first three-dimensional structure of a mammalian (GT29) sialyltransferase has, for the first time, allowed quantitative structure/function analyses to be performed, thereby providing reliable insights into the roles of key active site amino acids. Kinetic analyses of mutants of ST3Gal-I, in conjunction with structural studies, have confirmed the mechanistic roles of His302 and His319 as general acid and base catalysts, respectively, and have quantitated other interactions with the cytosine monophosphate-N-acetyl β-neuraminic acid donor substrate.

Structure and mechanism of the lipooligosaccharide sialyltransferase from Neisseria meningitidis.

The first x-ray crystallographic structure of a CAZY family-52 glycosyltransferase, that of the membrane associated α2,3/α2,6 lipooligosaccharide sialyltransferase from Neisseria meningitidis serotype L1 (NST), has been solved to 1.95 Å resolution. The structure of NST adopts a GT-B-fold common with other glycosyltransferase (GT) families but exhibits a novel domain swap of the N-terminal 130 residues to create a functional homodimeric form not observed in any other class to date.

Structural insight into mammalian sialyltransferases.

Mammalian cell surfaces are modified by complex arrays of glycoproteins, glycolipids and polysaccharides, many of which terminate in sialic acid and have central roles in essential processes including cell recognition, adhesion and immunogenicity. Sialylation of glycoconjugates is performed by a set of sequence-related enzymes known as sialyltransferases (STs). Here we present the crystal structure of a mammalian ST, porcine ST3Gal-I, providing a structural basis for understanding the mechanism and specificity of these enzymes and for the design of selective inhibitors.

Structural and kinetic characterizations of the polysialic acid O-acetyltransferase OatWY from Neisseria meningitidis.

The neuroinvasive pathogen Neisseria meningitidis has 13 capsular serogroups, but the majority of disease is caused by only 5 of these. Groups B, C, Y, and W-135 all display a polymeric sialic acid-containing capsule that provides a means for the bacteria to evade the immune response during infection by mimicking host sialic acid-containing cell surface structures. These capsules in serogroups C, Y, and W-135 can be further acetylated by a sialic acid-specific O-acetyltransferase, a modification that correlates with decreased immunoreactivity and increased virulence.

Benzofuran-derived cyclic beta-amino acid scaffold for building a diverse set of flavonoid-like probes and the discovery of a cell motility inhibitor.

We report here a practical, enantioselective synthesis of benzofuran-derived, cyclic trans-beta-amino acid scaffold. In two cases, tricyclic derivatives having six- and eight-membered unsaturated lactams were obtained from this versatile scaffold. To explore the biological applications, these compounds were subjected to cell-based assays, using NIH3T3 mouse cells to examine their potency as cell motility inhibitors and identified 18 as a potent cell motility inhibitor (IC50 approximately 40 microM in chamber cell migration assay).

Synthesis and bioorthogonal coupling chemistry of a novel cyclopentenone-containing unnatural tyrosine analogue.

Herein we report the synthesis of a novel amino acid with orthogonal functionality to the natural amino acid side chains. Tyrosine was O-alkylated with a cyclic 5-membered alpha,beta-unsaturated ketone ring (5). We have established that this amino acid analogue can undergo cycloaddition reactions in aqueous media with in situ generated nitrones.

A small-molecule probe for hepatitis C virus replication that blocks protein folding.

The hepatitis C virus (HCV) is a growing global health problem. Small molecules that interfere with host-viral interactions can serve as powerful tools for elucidating the molecular mechanisms of pathogenesis and defining new strategies for therapeutic development. Using a cell-based screen involving subgenomic HCV replicons, we identified the ability of 18 different abscisic acid (ABA) analogs, originally developed as plant growth regulators, to inhibit HCV replication.

Peroxisome proliferator-activated receptor alpha antagonism inhibits hepatitis C virus replication.

Hepatitis C virus (HCV) is a global health problem and a leading cause of liver disease. Here, we demonstrate that the replication of HCV replicon RNA in Huh-7 cells is inhibited by a peroxisome proliferator-activated receptor (PPAR) antagonist, 2-chloro-5-nitro-N-(pyridyl)benzamide (BA). Downregulation of PPARgamma with RNA interference approaches had no effect on HCV replication in Huh-7 cells, whereas PPARalpha downregulation inhibited HCV replication.

Exploring new chemical space by stereocontrolled diversity-oriented synthesis.

Natural products that act as highly specific, small-molecule protein-binding agents and as modulators of protein-protein interactions are highly complex and exhibit functional groups with three-dimensional and stereochemical diversity. The complex three-dimensional display of chiral functional groups appears to be crucial for exhibiting specificity in protein binding and in differentiating between closely related proteins. The development of methods that allow a high-throughput access to three-dimensional, skelatally complex, polycyclic compounds having few asymmetric diversity sites is essential and a highly challenging task.