<Superscript>13</Superscript>C APSY-NMR for sequential assignment of intrinsically disordered proteins

The increasingly recognized biological relevance of intrinsically disordered proteins requires a continuous expansion of the tools for their characterization via NMR spectroscopy, the only technique so far able to provide atomic-resolution information on these highly mobile macromolecules. Here we present the implementation of projection spectroscopy in ¹³C-direct detected NMR experiments to achieve the sequence specific assignment of IDPs. The approach was used to obtain the complete backbone assignment at high temperature of α-synuclein, a paradigmatic intrinsically disordered protein.     

Tandemly repeated satellite DNA ofDolichopoda schiavazzii: A test for models on the evolution of highly repetitive DNA

The evolutionary relationships among the Carnivora were studied in a phylogenetic analysis based on the complete mitochondrial cytochromeb gene. The study, which addressed primarily the relationships among the Caniformia, included 4 feliform and 26 caniform species, with 9 pinnipeds. The analysis identified five caniform clades: Canidae, Ailuridae (with the monotypic lesser panda), Musteloidea (Mustelidae+Procyonidae), Ursidae (including the giant panda), and Pinnipedia. The closest relatives of the Pinnipedia among terrestrial caniforms were not identified conclusively. Our analysis shows that the skunks are only distantly related to remaining mustelids (Mustelidae sensu stricto) and that the family Mustelidae, including the skunks, is paraphyletic. The relationship among the five caniform clades was unresolved, suggesting an evolutionary separation within a relatively short period of time. Based on distance values, we propose that this primary diversification took place 45 million years ago.     

Conformational studies of heterochiral peptides with diastereoisomeric residues: Crystal and molecular structures of linear dipeptides derived from leucine,isoleucine, and allo-isoleucine

The x-ray diffraction analyses of three N- and C-terminally blocked L , D dipeptides, namely t-Boc-D -Leu-L -Leu-OMe ( 1 ), t-Boc-L -Ile-D -alle-OMe ( 2 ), and t-Boc-D -aIle-L -Ile-OMe (3) containing enantiomeric or diastereomeric amino acid residues have been carried out. The structures were determined by direct methods and refined anisotropically to final R factors of 0.077. 0.058. and 0.072 for ( 1 ) ( 2 ) and ( 3 ), respectively. Peptides 1–3 all assume a similar U-shaped structure with ? and ψ torsion angles cosrresponding to one of the possible calculated minimum energy regions (regions E and G for L residues, and F*. D* and H* for D residues). The peptide backbones of 1-3 are almost super-imposable [provided that the appropriate inversion of the chiral centers of ( 2 ) is made]. Side-chain conformations of Leu residues in peptide ( 1 ) are g^? (tg^?) for the L -Leu residue and the mirrored g⁺ (tg⁺) for the D -Leu residue; however, in peptides ( 2 ) and ( 3 ) the conformations of the isoconfiguralional side chains of the Ile or allo-Ile residues are (g^?t) t and (tg⁺) tfor the L -Ile and the D -allo-Ile moieties, respectively. In all cases, these conformations correspond to the more populated conformers of β-branched residues statistically found in crystal structures of small peptides. The results seem to indicate that, at least in short peptides with enantiomeric or diastereoisomeric residues, the change in chirality in the main-chain atoms perturbs the backbone conformation to a lesser extent and the side chain conformation to a greater extent. © 1995 John Wiley & Sons, Inc.     

Reproductive features of homospecific hybridogenetically-derived stick insects suggest how unisexuals can evolve

Hybridogenetic reproduction has been demonstrated in both vertebrate and invertebrate unisexual hybrids. Its most peculiar feature is the transmission to the progeny of one invariant genome (hemiclone) through the egg and the replacement of the other by host fathering males. Bacillus hybridogens are the only known example of hemiclonal invertebrates; their comparison to Poeciliopsis and Rana systems helps in understanding peculiar and shared features of vertebrate and insect hybridogenesis. In P. monacha-lucida, the experimental production of non-hybrid progeny through the reunion of the maternal hemiclone with a homospecific paternal genome provided by males of the maternal ancestor leads to inviable or severely impaired sterile specimens, whereas in Rana esculenta viable offspring are the rule. The comparable synthetic B. rossius progeny (Rr) embodying the maternal R hemiclone and a paternal r haploset, appear perfectly viable and fertile, clearly demonstrating compatibility between the two homospecific genomes, and also supporting a lack of deterioration of the R hemiclone. This condition can be ascribed to the recent origin of the hemiclones, and also to the absence of lethal recessives, owing to their most likely derivation from an automictic doubling in the parthenogenetic mechanisms of the maternal ancestor. However, the hybridogenetic system breaks down in the gamete production of the majority of Rr females, since normal allele segregation also occurs in their progeny. These reproductive modes suggest a likely evolutionary dynamic for newly originated hybridogens: to achieve stability, an interruption of reproductive interactions with the maternal ancestor seems necessary. In stick insects, this constraint appears to be fulfilled in both areas of sympatry. The microevolutionary pathway suggested by the ecological scenario also supports the possibility that a shift of hemiclonal stick insect strains to clonality has occurred.     

Multiple synthesis by the multipin method as a methodological tool

The multipin method of peptide synthesis is demonstrated as a potent methodological tool, where large numbers of comparative studies can be performed concurrently. Two studies are presented. In each study, the test peptides were simultaneously synthesized, and the products examined by high throughput ion spray mass spectrometry and reverse-phase HPLC. In the first study, comprising 24 experiments, peptides 1 (AELFSTHYLAFKEDYSQ-NH₂) and 2 (LKDFRVYFREGRDQLWKGPG-NH₂) were prepared using Fmoc-Axx/BOP/HOBt/NMM (100: 100: 100: 150 mM ) and Fmoc-Axx/HATU/HOAt/NMM (100: 100: 100: 150 mM ) with 60.90 and 120 min coupling times. The two reagent combinations were found to give comparable results. The second study compared the N-terminal coupling of Fmoc-Asn-OH, Fmoc-Asn(Mbh)-OH, Fmoc-Asn(Mtt)-OH, Fmoc-Asn(Tmob)-OH and Fmoc-Asn(Trt)-OH in the synthesis of seven test peptides: 3 , NVQAAIDYIG-cyclo(Kp); 4 , NTVQAAIDYIG-cyclo(KF); 5 , NRVYVHPFNL; 6 , NRVYVHPFHL: 7 , NEAYVHDAPVRSLN: 8 , NQLVVPSEGLYLIYSQVLFK. 9 , NPNANPNANPNA. A total of 33 experiments were performed. Peptides 3 and 4 were selected to highlight the effect of steric bulk of each Asn derivative on coupling efficiency. Reagent efficiency, as measured by target peptide purity, was as follows: Fmoc-Asn(Tmob)-OH > Fmoc-Asn-OH > Fmoc-Asn(Mtt)-OH = Fmoc-Asn(Trt)-OH > Fmoc-Asn(Mbh)-OH.     

The development of benthonic phytocoenosis on artificial substrates in the ticino river

Summary Changes in the taxonomic composition, chlorophyll a concentration, dry weight, percentage organic carbon and nitrogen and several indices of diversity, including Margalef's index were followed during the development of phytobenthonic communities on glass slides. These data suggest that, in this environment, the algal community resembles the nearby natural community after 4 weeks. The taxonomic development can be divided into two phases. During the first 2 weeks the phytocoenosis is dominated by a rather diverse and variable diatom assemblage. Later Cyanophyta dominate. The diversity decreases during colonization as reflected by all indices considered.     

Blood groups of crab-eating macaques (Macaca fascicularis) demonstrated by isoimmune rhesus monkey (Macaca mulatta) sera.

Twenty-one isoimmune sera produced in rhesus monkey (Macaca mulatta) containing type-specific antibodies for simian-type red cell antigens were tested for their cross-reactivity with red cells from crab-eating macaques (M. fascicularis). The majority of the antisera gave cross-reactions determining polymorphisms in the red cells of crab-eating macaques, homologous to those of rhesus monkeys. These results attest to the close taxonomic realationship between the two species of macaques, and have the practical implication that isoimmune sera produced for blood typing can also be used for typing red cells from related species, as has been also observed in studies on apes.