Purine Nucleoside Phosphorylase Deficiency: A Mutation Update

Purine nucleoside phosphorylase (PNPase) deficiency is an autosomal recessive disorder affecting purine degradation and salvage pathways. Clinically, patients typically present with severe immunodeficiency, neurological dysfunction, and autoimmunity. Biochemically, PNPase deficiency may be suspected in the presence of hypouricemia. We report biochemical and genetic data on a cohort of seven patients from six families identified as PNPase deficient. In all patients, inosine, deoxyinosine, guanosine, and deoxyguanosine were elevated in urine, and mutation analysis revealed seven different mutations of which three were novel. The mutation c.770A>G resulted in the substitution p.His257Arg. A second novel mutation c.257A>G (p.His86Arg) was identified in two siblings and a third novel mutation, c.199C>T (p.Arg67X), was found in a 2-year-old female with delayed motor milestones and recurrent respiratory infections. A review of the literature identified 67 cases of PNPase deficiency from 49 families, including the cases from our own laboratory. PNPase deficiency was confirmed in 30 patients by genotyping and 24 disease causing mutations, including the three novel mutations described in this paper, have been reported to date. In five of the seven patients, plasma uric acid was found to be within the pediatric normal range, suggesting that PNPase deficiency should not be ruled out in the absence of hypouricemia.     

The immune evasion protein Sbi of Staphylococcus aureus occurs both extracellularly and anchored to the cell envelope by binding lipoteichoic acid

The Sbi protein of Staphylococcus aureus comprises two IgG‐binding domains similar to those of protein A and a region that triggers the activation of complement C3. Sbi is expressed on the cell surface but its C‐terminal domain lacks motifs associated with wall or membrane anchoring of proteins in Gram‐positive bacteria. Cell‐associated Sbi fractionates with the cytoplasmic membrane and is not solubilized during protoplast formation. S. aureus expressing Sbi truncates of the C‐terminal Y domain allowed identification of residues that are required for association of Sbi with the membrane. Recombinant Sbi bound to purified cytoplasmic membrane material in vitro and to purified lipoteichoic acid. This explains how Sbi partitions with the membrane in fractionation experiments yet is partially exposed on the cell surface. An LTA‐defective mutant of S. aureus had reduced levels of Sbi in the cytoplasmic membrane.     

Significance of nucleotide sequence alignments: a method for random sequence permutation that preserves dinucleotide and codon usage

The similarity of two nucleotide sequences is often expressed in terms of evolutionary distance, a measure of the amount of change needed to transform one sequence into the other. Given two sequences with a small distance between them, can their similarity be explained by their base composition alone? The nucleotide order of these sequences contributes to their similarity if the distance is much smaller than their average permutation distance, which is obtained by calculating the distances for many random permutations of these sequences. To determine whether their similarity can be explained by their dinucleotide and codon usage, random sequences must be chosen from the set of permuted sequences that preserve dinucleotide and codon usage. The problem of choosing random dinucleotide and codon-preserving permutations can be expressed in the language of graph theory as the problem of generating random Eulerian walks on a directed multigraph. An efficient algorithm for generating such walks is described. This algorithm can be used to choose random sequence permutations that preserve (1) dinucleotide usage, (2) dinucleotide and trinucleotide usage, or (3) dinucleotide and codon usage. For example, the similarity of two 60-nucleotide DNA segments from the human beta-1 interferon gene (nucleotides 196-255 and 499-558) is not just the result of their nonrandom dinucleotide and codon usage.      

Stereospecific pharmacokinetics and toxicodynamics of ketorolac after oral administration of the racemate and optically pure enantiomers to the rat

To determine the stereospecific pharmacokinetics and gastrointestinal permeability (GI) changes (surrogate measures of toxicity) in the rat following oral administration of S, R, and racemic ketorolac (KT), optically pure enantiomers (S and R 2.5 mg/kg), and racemic KT (5 mg/kg) were administered orally to male Sprague-Dawley rats and plasma samples were collected for 6 h post-dose for pharmacokinetic assessments. KT-induced changes in GI permeability were assessed using sucrose and 51Cr-EDTA as markers of gastroduodenal and distal intestinal permeability, respectively. After the racemate, R-KT was predominant in plasma (AUC S/R, 0.45). No significant differences in pharmacokinetic indices were evident following administration of the racemate as compared with individual enantiomers. In plasma, there was only negligible S-KT after administration of R-KT. After S-KT, on the other hand, AUC of R-KT was found to be 6.7% of that of S-KT. Both permeability markers showed considerable interanimal variability. Gastroduodenal permeability was significantly increased from baseline by the racemate but not by either of the two enantiomers administered alone. Permeability to 51Cr-EDTA was not significantly increased above baseline for any of the treatments. The plasma concentration of R-KT found after administration of S-KT may be from the < 2% chiral impurity which appears magnified due to its slower clearance as compared with its antipode. There is no evidence of a pharmacokinetic interaction between the enantiomers. Since 2.5 mg/kg S-KT is somewhat less toxic on the gastroduodenum than 5 mg/kg racemate, it may be a safer alternative to the latter, at least in the rat model.     

Galápagos and Californian sea lions are separate species: Genetic analysis of the genus Zalophus and its implications for conservation management

The climatic cycles with subsequent glacial and intergalcial periods have had a great impact on the distribution and evolution of species. Using genetic analytical tools considerably increased our understanding of these processes. In this review I therefore give an overview of the molecular biogeography of Europe. For means of simplification, I distinguish between three major biogeographical entities: (i) "Mediterranean" with Mediterranean differentiation and dispersal centres, (ii) "Continental" with extra-Mediterranean centres and (iii) "Alpine" and/or "Arctic" with recent alpine and/or arctic distribution patterns. These different molecular biogeographical patterns are presented using actual examples. Many "Mediterranean" species are differentiated into three major European genetic lineages, which are due to glacial isolation in the three major Mediterranean peninsulas. Postglacial expansion in this group of species is mostly influenced by the barriers of the Pyrenees and the Alps with four resulting main patterns of postglacial range expansions. However, some cases are known with less than one genetic lineage per Mediterranean peninsula on the one hand, and others with a considerable genetic substructure within each of the Mediterranean peninsulas, Asia Minor and the Maghreb. These structures within the Mediterranean sub-centres are often rather strong and in several cases even predate the Pleistocene. For the "Continental" species, it could be shown that the formerly supposed postglacial spread from eastern Palearctic expansion centres is mostly not applicable. Quite the contrary, most of these species apparently had extra-Mediterranean centres of survival in Europe with special importance of the perialpine regions, the Carpathian Basin and parts of the Balkan Peninsula. In the group of "Alpine" and/or "Arctic" species, several molecular biogeographical patterns have been found, which support and improve the postulates based on distribution patterns and pollen records. Thus, genetic studies support the strong linkage between southwestern Alps and Pyrenees, northeastern Alps and Carpathians as well as southeastern Alps and the Dinaric mountain systems, hereby allowing conclusions on the glacial distribution patterns of these species. Furthermore, genetic analyses of arctic-alpine disjunct species support their broad distribution in the periglacial areas at least during the last glacial period. The detailed understanding of the different phylogeographical structures is essential for the management of the different evolutionary significant units of species and the conservation of their entire genetic diversity. Furthermore, the distribution of genetic diversity due to biogeographical reasons helps understanding the differing regional vulnerabilities of extant populations.     

Problems in the measurement of zinc using heparin as an anticoagulant

Although several reports imply that anticoagulants and preservatives contain zinc, the quantity of zinc in heparin, if any, has not been documented. Zinc concentration was determined by flame atomic absorption spectroscopy in varying dilutions of multiple commercially obtained samples of purified sodium heparin N = 15 (microgram Zn/1000 Units heparin). Rubber stoppers of sterile heparin vials and of blood evacuation tubes were incubated in pre-analyzed water or saline on a mechanical shaker with fluid aliquots obtained up to 27 hours and analyzed for zinc content (microgram Zn/0.1 ml). Heparin, with contact or without contact with rubber stoppers, recorded identical zinc concentrations. Zinc concentrations varied from 0.222 +/- 0.01 (mean +/- SE) to 3.49 +/- 0.005 microgram Zn/1000 Units heparin. Leaching of zinc from rubber stoppers of vacutainer tubes (N = 9) was noted only with those containing known chelators of zinc. These results indicate that zinc is present in certain lots of sodium heparin and that caution must be exercised when reporting zinc concentrations of blood samples that contain sodium heparin as the anticoagulant.