Application of the imidate procedure to α-d-galactosyltion

Using the imidate procedure, 2,3,4,6-tetra-O-benzyl-1-O-(N-methylacetimidoyl)-β-d-galactopyranose was condensed with various monosaccharides to provide, in good yield and with high stereoselectivity, α-linked disaccharides.     

Amyloid kidney stones of uremic patients consist of beta 2-microglobulin fragments

Urinary stones with amyloid structure, obtained from uremic patients, were analyzed according to molecular weight, amino acid sequence, and antigenic content. A major protein of approximately 7 kD, designated AB protein, was isolated by size exclusion using HPLC in 60% formic acid. AB protein reacted in immunodiffusion only with an antiserum to beta 2-microglobulin, with beta 2m spurring over AB protein. N-terminal amino acid sequence analysis defined two fragments homologous to beta 2m. One fragment commenced with Ile at position 7 and the other with Ser at position 20, with a cleavage point subsequent to a lysyl residue in both. It is concluded that beta 2m is a precursor of urinary amyloid stones and intratubular concretions of patients with preterminal and terminal renal failure; limited proteolysis is involved in AB amyloid generation.     

Synthesis of M and N active glycopeptides. Part of the N-terminal region of human glycophorin A

The synthesis of two series of glycopeptides, part of the N-terminal region of human glycophorin A, was accomplished starting from derivatives ofO--d-galactopyranosyl-(1–3)-O-(2-acetamido-2-deoxy--d-galactopyranosyl)-l-serine and-l-threonine.     

MRS3 and MRS4, two suppressors of mtRNA splicing defects in yeast, are new members of the mitochondrial carrier family.

When present in high copy number plasmids, the nuclear genes MRS3 and MRS4 from Saccharomyces cerevisiae can suppress the mitochondrial RNA splicing defects of several mit- intron mutations. Both genes code for closely related proteins of about Mr 32,000; they are 73% identical. Sequence comparisons indicate that MRS3 and MRS4 may be related to the family of mitochondrial carrier proteins. Support for this notion comes from a structural analysis of these proteins. Like the ADP/ATP carrier protein (AAC), the mitochondrial phosphate carrier protein (PiC) and the uncoupling protein (UCP), the two MRS proteins have a tripartite structure; each of the three repeats consists of two hydrophobic domains that are flanked by specific amino acid residues. The spacing of these specific residues is identical in all domains of all proteins of the family, whereas spacing between the hydrophobic domains is variable. Like the AAC protein, the MRS3 and MRS4 proteins are imported into mitochondria in vitro and without proteolytic cleavage of a presequence and they are located in the inner mitochondrial membrane. In vivo studies support this mitochondrial localization of the MRS proteins. Overexpression of the MRS3 and MRS4 proteins causes a temperature-dependent petite phenotype; this is consistent with a mitochondrial function of these proteins. Disruption of these genes affected neither mitochondrial functions nor cellular viability. Their products thus have no essential function for mitochondrial biogenesis or for whole yeast cells that could not be taken over by other gene products. The findings are discussed in relation to possible functions of the MRS proteins in mitochondrial solute translocation and RNA splicing.     

The nuclear gene MRS2 is essential for the excision of group II introns from yeast mitochondrial transcripts in vivo.

RNA splicing defects in mitochondrial intron mutants can be suppressed by a high dosage of several proteins encoded by nuclear genes. In this study we report on the isolation, nucleotide sequence, and possible functions of the nuclear MRS2 gene. When present on high copy number plasmids, the MRS2 gene acts as a suppressor of various mitochondrial intron mutations, suggesting that the MRS2 protein functions as a splicing factor. This notion is supported by the observations that disruption of the single chromosomal copy of the MRS2 gene causes (i) a pet- phenotype and (ii) a block in mitochondrial RNA splicing of all four mitochondrial group II introns, some of which are efficiently self-splicing in vitro. In contrast, the five group I introns monitored here are excised from pre-mRNA in a MRS2-disrupted background although at reduced rates. So far the MRS2 gene product is unique in that it is essential for splicing of all four group II introns, but relatively unimportant for splicing of group I introns. In strains devoid of any mitochondrial introns the MRS2 gene disruption still causes a pet- phenotype and cytochrome deficiency, although the standard pattern of mitochondrial translation products is produced. Therefore, apart from RNA splicing, the absence of the MRS2 protein may disturb the assembly of mitochondrial membrane complexes.     

Oxidative stress‐induced renal telomere shortening as a mechanism of cyclosporine‐induced nephrotoxicity

Due to the association of oxidative stress and telomere shortening, it was aimed in the present study to investigate the possibility whether cyclosporine‐A exerts its nephrotoxic side effects via induction of oxidative stress‐induced renal telomere shortening and senescent phenotype in renal tissues of rats. Renal oxidative stress markers, 8‐hydroxydeoxyguanosine, malondialdehyde, and protein carbonyl groups were measured by standard methods. Telomere length and telomerase activity were also evaluated in kidney tissue samples. Results showed that cyclosporine‐A treatment significantly (P < 0.05) enhanced renal malondialdehyde, 8‐hydroxydeoxyguanosine, and protein carbonyl groups levels, decreased renal telomere length, and deteriorated renal function compared with the controls. Renal telomerase activity was not affected by cyclosporine‐A. Renal telomere length could be considered as an important parameter of both oxidative stress and kidney function. Telomere shortening and accelerated kidney aging may be caused by cyclosporine‐induced oxidative stress, indicating the potential mechanism of cyclosporine‐induced nephrotoxicity.     

Anthrax biosensor, protective antigen ion channel asymmetric blockade

The significant threat posed by biological agents (e.g. anthrax, tetanus, botulinum, and diphtheria toxins) (Inglesby, T. V., O'Toole, T., Henderson, D. A., Bartlett, J. G., Ascher, M. S., Eitzen, E., Friedlander, A. M., Gerberding, J., Hauer, J., Hughes, J., McDade, J., Osterholm, M. T., Parker, G., Perl, T. M., Russell, P. K., and Tonat, K. (2002) J. Am. Med. Assoc. 287, 2236-2252) requires innovative technologies and approaches to understand the mechanisms of toxin action and to develop better therapies. Anthrax toxins are formed from three proteins secreted by fully virulent Bacillus anthracis, protective antigen (PA, 83 kDa), lethal factor (LF, 90 kDa), and edema factor (EF, 89 kDa). Here we present electrophysiological measurements demonstrating that full-length LF and EF convert the current-voltage relationship of the heptameric PA63 ion channel from slightly nonlinear to highly rectifying and diode-like at pH 6.6. This effect provides a novel method for characterizing functional toxin interactions. The method confirms that a previously well characterized PA63 monoclonal antibody, which neutralizes anthrax lethal toxin in animals in vivo and in vitro, prevents the binding of LF to the PA63 pore. The technique can also detect the presence of anthrax lethal toxin complex from plasma of infected animals. The latter two results suggest the potential application of PA63 nanopore-based biosensors in anthrax therapeutics and diagnostics.     

Achillea filipendulina Lam.: Chemical Constituents and Antimicrobial Activities of Essential Oil of Stem,Leaf, and Flower

In this study, we extracted the essential oils of the stem, leaf, and flower of Achillea filipendulina, analyzed them, and studied their antibacterial properties. Of 16, 53, and 35 compounds identified in the stem, leaf, and flowers, respectively, only five are present in all three segments of the plant. The essential oil of the stem was mainly composed of neryl acetate, spathulenol, carvacrol, santolina alcohol, and trans‐caryophyllene oxide. However, the main identified components of leaf were 1,8‐cineole, camphor, ascaridole, trans‐isoascaridole, and piperitone oxide and the main components of the flower oil were ascaridole, trans‐isoascaridole, 1,8‐cineole, p‐cymene, and camphor. The extracted oil from different segments demonstrated varying antibacterial properties against both Gram‐positive and Gram‐negative bacteria, demonstrated by disk, minimum inhibitory concentration, and minimum bactericidal concentration methods. These suggest that the application of all segments of aerial parts of A. filipendulina may have a better therapeutic effect in fighting pathogenic systems.     

SANS (USH1G) regulates pre-mRNA splicing by mediating the intra-nuclear transfer of tri-snRNP complexes

Splicing is catalyzed by the spliceosome, a compositionally dynamic complex assembled stepwise on pre-mRNA. We reveal links between splicing machinery components and the intrinsically disordered ciliopathy protein SANS. Pathogenic mutations in SANS/USH1G lead to Usher syndrome—the most common cause of deaf-blindness. Previously, SANS was shown to function only in the cytosol and primary cilia. Here, we have uncovered molecular links between SANS and pre-mRNA splicing catalyzed by the spliceosome in the nucleus. We show that SANS is found in Cajal bodies and nuclear speckles, where it interacts with components of spliceosomal sub-complexes such as SF3B1 and the large splicing cofactor SON but also with PRPFs and snRNAs related to the tri-snRNP complex. SANS is required for the transfer of tri-snRNPs between Cajal bodies and nuclear speckles for spliceosome assembly and may also participate in snRNP recycling back to Cajal bodies. SANS depletion alters the kinetics of spliceosome assembly, leading to accumulation of complex A. SANS deficiency and USH1G pathogenic mutations affects splicing of genes related to cell proliferation and human Usher syndrome. Thus, we provide the first evidence that splicing dysregulation may participate in the pathophysiology of Usher syndrome.