Shape of the pelvis and posture of the hindlimbs inPlateosaurus

The pelvis ofPlateosaurus is examined from a biomechanical point of view. The shape of the acetabulum in particular is analysed in order to determine the range of possible directions of the forces exchanged between femur and pelvis. These forces must have been more or less confined to a sagittal plane. From a quasi-static analysis under consideration of the major hip muscles ofPlateosaurus, a nearly but not fully extended posture of the hindlimbs can be deduced. The hip joints ofPlateosaurus and probably of some other dinosaurs with a narrow biacetabular width were balanced rather by adducting than by abducting muscles.     

Anchor-linked intermediates in peptide amide synthesis are caused by dimeric anchors on the solid supports

Cleavage and kinetic studies have been carried out using commercially obtained H-Tyr(tBu)-5-(4′-aminomethyl-3′,5′-dimethoxyphenoxy)valeric acid-TentaGelS (H-Tyr(tBu)-4-ADPV-TentaGelS) and H-Tyr (tBu)-4-ADPV-Ala-aminomethyl-resin (H-Tyr(tBu)-4-ADPV-AM-resin) prepared from commercially available resin and loaded with commercially available Fmoc-4-ADPV-OH amide anchor. Cleavage with pure trifluoroacetic acid (TFA) gave the intermediate H-Tyr-4-ADPV-NH₂, which was then degraded to H-Tyr-NH₂, and cleavage with TFA/dichloromethane (1:9) yielded H-Tyr-4-ADPV-NH₂ which could be isolated in preparative amounts. Cleavage reactions with ¹⁵N-labelled H-Ala-4-ADPV-[¹⁵N]-Gly-AM-resin yielded the intermediate H-Ala-4-ADPV-NH₂, which contained no ¹⁵N as demonstrated by ¹H-NMR. The analysis of the commercial Fmoc-4-ADPV-OH amide anchor showed the presence of Fmoc-4-ADPV-4-ADPV-OH as an impurity in high amounts. This dimeric anchor molecule is the cause of formation of the anchor-linked peptide intermediate obtained during the cleavage from the resin. The particularly high acid-lability of the amide bond between the two ADPV moieties was utilized to synthesize sidechain and C-terminally 4-ADPV protected pentagastrin on a double-anchor resin, and to cleave it using 5% trifluoroacetic acid in dichloromethane. This method may offer a new way for the synthesis of protected peptide amides with improved solubility to be used in fragment condensation.     

Characterization of mutations on the rare duplicated C4/CYP21 haplotype in steroid 21-hydroxylase deficiency

We have defined the mutations causing congenital adrenal hyperplasia in three Swedish patients carrying a rare haplotype containing two mutated steroid 21-hydroxylase genes (CYP21) in addition to one pseudogene (CYP21P). The presence of such haplotypes complicates genetic diagnosis and screening of mutations in 21-hydroxylase deficiency, and we show how these genotypes can be resolved by amplification and analysis of each gene separately. In all cases, the rare haplotype carried the same combination of disease-causing mutations; one of the genes had the splice mutation at base 659 in intron 2, and the other had the nonsense mutation at base 1999 in exon 8 (CAG to TAG). We have thus characterized the most common haplotype containing duplicated CYP21 genes. The frequency of this haplotype is low, and if additional such haplotypes are present, they are rare in this population.     

G2 phase cell cycle disturbance as a manifestation of genetic cell damage

The predominant cell cycle change induced by X-rays and clastogens in peripheral blood mononuclear cells is the accumulation of cells in the G2 phase of the cell cycle. We show that this accumulation consists of cells that are either delayed or arrested within the G2 phase. Since both X-rays and DNA crosslinking chemicals are known to damage DNA, the G2 phase inhibition caused by these agents is thought to be one of the primary manifestations of (unrepaired) DNA damage. This interpretation is supported by two additional findings. (1) Older individuals have elevated baseline levels of mononuclear blood cells that are delayed and/or arrested in the G2 phase of the cell cycle. This coincides with the increased chromosomal breakage rates reported for older individuals. (2) Irrespective of their age, individuals with inherited genetic instability syndromes (such as Fanconi anemia and Bloom syndrome) exhibit elevated G2 phase cell fractions. We show that the method used to detect such induced or spontaneous cell cycle changes, viz. BrdU-Hoechst flow cytometry, is a rapid and highly sensitive technique for the assessment of genetic cell damage.Dedicated to Professor Ulrich Wolf on the occasion of his 60th birthday     

Tight linkage between the Beckwith-Wiedemann syndrome and a microsatellite marker for the TH locus

The Beckwith-Wiedemann syndrome (BWS) is characterized by somatic overgrowth, developmental anomalies, and proneness to embryonic tumor development. The majority of cases are sporadic, but several families with an autosomal dominant mode of inheritance with variable expression and reduced penetrance have been described. In three such families, BWS has been linked to DNA markers for the insulin gene (INS) and H-ras on chromosome band 11p15. Two additional families with inherited BWS are described here. Linkage analysis has been performed with a highly informative marker for the tyrosine hydroxylase (TH) locus within the INS-IGF2 (insulin-like growth factor II)-TH gene cluser and confirms the previous observed linkage to this region (lod score 2.16 at = 0). Linkage analysis to TH provides a basis for informed genetic counselling and carrier detection in the hereditary form of the syndrome. Based on the hypothesis that IGF2 may be a candidate gene for BWS, we screened for mutations in the coding exons 7 and 9, but found no abnormalities in 5 unrelated BWS cases.     

New method for peptide purification based on selective removal of truncation peptide impurities after SPPS with orthogonal capping

Peptide purification by high-performance liquid chromatography (HPLC) is associated with high solvent consumption, relatively large effort and lack of efficient parallelization. As an alternative, many catch-and-release (c&r) purification methods have been developed over the last decades to enable the efficient parallel purification of peptides originating from solid-phase peptide synthesis (SPPS). However, with one exception, none of the c&r systems has been widely established in industry and academia until today. Herein, we present an entirely new chromatography-free purification concept for peptides synthesized on a solid support, termed reactive capping purification (RCP). The RCP method relies on the capping of truncation peptides arising from incomplete coupling of amino acids during SPPS with a reactive tag. The reactive tag contains a masked functionality that, upon liberation during cleavage from the resin, enables straightforward purification of the peptide by incubation with a resin-bound reactive moiety. In this work, two different reactive tags based on masked thiols were developed. Capping with these reactive tags during SPPS led to effective modification of truncated sequences and subsequent removal of the latter by chemoselective reaction with a maleimide-functionalized solid support. By introducing a suitable protecting group strategy, the thiol-based RCP method described here could also be successfully applied to a thiol-containing peptide. Finally, the purification of a 15-meric peptide by the RCP method was demonstrated. The developed method has low solvent consumption, has the potential for efficient parallelization, uses readily available reagents, and is experimentally simple to perform.     

Ornibactins—a new family of siderophores from Pseudomonas

Novel linear hydroxamate/hydroxycarboxylate siderophores from strains of Pseudomonas cepacia were isolated and named ornibactins. The ornibactins represent modified tetrapeptide siderophores, possessing the sequence l-Orn¹(N -OH, N -acyl)-d-threo-Asp(-OH)-l-Ser-l-Orn⁴(N -OH, N -formyl)-1,4-diaminobutane. The N -acyl groups of Orn¹(N -OH, N -acyl) may vary and represent the three acids 3-hydroxybutanoic acid, 3-hydroxyhexanoic acid and 3-hydroxyoctanoic acid, leading to a mixture of three different ornibactins, designated according to their acyl chain length as ornibactin-C4, ornibactin-C6 and ornibactin-C8. Each of the siderophores is accompanied by a small amount of a more hydrophilic component with a 16 a.m.u. higher mass. The structure elucidation was based on results from gas chromatography amino acid analysis, electrospray mass spectrometry, and one- and two-dimensional nuclear magnetic resonance techniques.     

Diversity of Siphonaria Sowerby I, 1823 (Gastropoda,Siphonariidae) in the Seychelles Bank and beyond

Molecular phylogenetic studies have shown that the characters of the reduced shell of the false limpets of the genus Siphonaria Sowerby I, 1823 are highly variable and often insufficient for species delimitation. The taxonomy and distribution of Siphonaria in the Indian Ocean are poorly known. We sampled Siphonaria in the Seychelles Bank to check the occurrence of recorded species using DNA sequences and to study the paths through which Siphonaria species have colonised the Seychelles Bank by reconstructing their phylogenetic relationships. Analyses of a dataset comprising 16 S rRNA gene sequences of 33 specimens from the Seychelles Bank and 300 additional Siphonaria sequences from other regions from GenBank with various methods for species delimitation resulted in 19–102 primary species hypotheses. Assemble Species by Automatic Partitioning provided a conservative estimate of the species number (42) in which several indisputable species were lumped. The results of Automatic Barcode Gap Discovery depended strongly on the assumed prior maximum intraspecific divergence, whereas the tree-based methods Generalised Mixed Yule Coalescent and Poisson Tree Processes resulted in high overestimates. The specimens from the Seychelles Bank represent three clades, belonging to the Siphonaria ‘atra’ group, the Siphonaria ‘normalis’ group and a possibly undescribed species recorded previously only from Hainan. At least two of the three species recorded from the Seychelles Bank came from the east, i.e., from the Coral Triangle in the Indo-Australian Archipelago, the region with the highest marine biodiversity worldwide. A major transport mechanism across the Indian Ocean was probably the South Equatorial Current.