Spectroscopic studies of structural changes in two beta-sheet-forming peptides show an ensemble of structures that unfold noncooperatively

A characterization of the conformation and stability of model peptide systems that form beta-sheets in aqueous solutions is considerably important in gaining insights into the mechanism of beta-sheet formation in proteins. We have characterized the conformation and equilibrium folding and unfolding of two 20-residue peptides whose NMR spectra suggest a three-stranded beta-sheet topology in aqueous solution: Betanova [Kortemme, T., Ramirez-Alvarado, M., and Serrano, L. (1998) Science 281, 253-256] and (D)P(D)P with d-Pro-Gly segments at the turns [Schenck, H. L., and Gellman, S. H. (1998) J. Am. Chem. Soc. 120, 4869-4870]. Both circular dichroism (CD) and infrared measurements indicate only 20-26% beta-sheet-like structure at 5 degrees C for Betanova and 42-59% beta-sheet for (D)P(D)P. For both peptides, the CD and infrared spectra change nearly linearly with increasing temperatures (or urea concentrations) and lack a sigmoidal signature characteristic of cooperative unfolding. Fluorescence resonance energy transfer (FRET) measurements between donor and acceptor molecules attached to the two ends confirm that Betanova is largely unstructured even at 10 degrees C; the average end-to-end distance estimated from FRET is closer to that of a random coil than a structured beta-sheet. In (D)P(D)P, the FRET results indicate a more compact structure that remains compact even at high temperatures (approximately 80 degrees C) or high urea concentrations (approximately 8 M). These results indicate that both these peptides access an ensemble of conformations at all temperatures or denaturant concentrations, with no significant free energy barrier separating the "folded" and "unfolded" conformations.     

Response of the NAD(P)H-oxidising flavohaemoglobin (Hmp) to prolonged oxidative stress and implications for its physiological role in Escherichia coli

The Escherichia coli flavohaemoglobin (Hmp) has a globin-like N-terminal domain and a ferredoxin-NADP-reductase-like C-terminal domain. We show here that purified Hmp oxidises both NADH and NADPH with K _m values of 1.8 and 19.6 μM, respectively. Prolonged incubation of a hmp-lacZ fusion strain with the redox cycling agent paraquat resulted in a 28-fold induction of hmp gene expression, nearly 3-fold higher than after short periods of exposure. A strain overproducing Hmp was significantly more sensitive to paraquat than was the wild-type strain but, in vitro, purified Hmp was not an effective NADPH-paraquat diaphorase. Prolonged incubation of a wild-type strain with paraquat increased intracellular Hmp to spectrally detectable levels.     

India and the Muslim Punjab: a unified approach to South Asian kinship

The comparison of kinship of different South Asian regions with the Muslim Punjab reveals basic invariants, thus denying a fundamental division between the Dravidian South and the Indo-Aryan North, though concepts of kinship reveal themselves in different media: Muslim Punjabi kinship is expressed primarily in gift exchange, but exhibits characteristics commonly associated with the South Indian terminology. The present article argues for an approach which takes meaning embedded in the 'act' or 'person' as non-separable from 'culture', and grasps kinship as structuring the dynamic relation between self and other, regarded as ontological categories.  

Résumé

La comparaison de la parenté entre les différentes régions du sud de l'Asie et le Pendjab musulman révèle des constantes fondamentales et remet ainsi en question la division de principe entre le Sud dravidien et le Nord indo-aryen, bien que les concepts de parenté trouvent des expressions différentes : dans le Pendjab musulman, la parenté s'exprime principalement à travers l'échange de dons mais présente des caractéristiques habituellement associées à la terminologie du sud de l'Inde. Le présent article plaide en faveur d'une approche considérant la signification de « l'acte >> ou de la « personne >> en tant qu'entité indissociable de la « culture >>, et aborde la parenté comme un moyen de structurer la relation dynamique entre soi-même et l'autre, considérés comme des catégories ontologiques.     

Effect of dinoprost and cloprostenol on serum nitric oxide and corpus luteum blood flow during luteolysis in ewes

In this study we compared the effect of dinoprost and cloprostenol on changes of corpus luteum blood flow during luteolysis. Ten nonlactating cyclic ewes were synchronized with double PGF_2α injections 11 days apart. At Day 10, the animals were classified into 2 groups and received the third dose of PGF_2α after confirmation of the presence of a mature CL. The first group received (12.5 mg/im) dinoprost and the second group received (250 μg/im) cloprostenol. A color Doppler ultrasound scan was performed by the same operator according to the following timeline: 0, 0.5, 1, 2, 4, 6, 12, and 24 hours, then every 24 hours until Day 4). The size, morphology, and blood flow of the CL was evaluated during the regression. The results showed that regression of the CL did not differ between the dinoprost and cloprostenol groups. There was no significant effect on diameter of the CL in both groups, though the size of the CL decreased gradually and slowly. Pretreatment progesterone concentration did not differ between groups. The results showed that the nitric oxide level was significantly increased within half an hour after the dinoprost treatment, and was significantly decreased in the cloprostenol group after half an hour. The blood velocity was increased significantly half an hour after the dinoprost treatment and it was decreased in the cloprostenol-treated group. In conclusion, both cloprostenol and dinoprost affect CL by controlling the nitric oxide level and blood supply of the CL via different mechanisms to induce luteolysis.     

DNA typing of Pakistani cattle breeds Tharparkar and Red Sindhi by microsatellite markers

Microsatellite markers are used for any individual identity and breed characterization in animals that is an efficient and successful way of investigation. They are used for multiple purposes as genetic detectors including, rapid mutation rate, high level of polymorphism, and range of variety of microsatellite markers available. A panel of 19 microsatellite markers was developed for breed characterization in Tharparkar and Red Sindhi breeds of cattle in Pakistan. Forty four blood samples of cattle (each breed) were collected from Department of Livestock Management, Sindh Agriculture University, Tandojam, Tando Qaiser, Tharparkar Cattle Farm Nabi sar Road, Umer Kot, Sindh, and Govt. Red Sindhi Cattle Breeding Farm, Tando Muhammad Khan Pakistan. Breed characterization was 100% successful. Average PIC, He and Power of Exclusion values were found to be 0.91, 0.62 and 13.28, respectively. Pattern of allelic frequencies of most of the microsatellite markers were clearly distinct between two breeds. As a result of present study a reliable, efficient and very informative panel of microsatellite markers was successfully developed which was capable to interpret individual identity, forensic cases and breed characterization in cattle. This facility is ready to be provided to local cattle breeder at commercial level for DNA testing of cattle. This study will also be highly helpful for breed conservation of cattle. In addition this study can also become a basis to open up new disciplines of animal forensics in Pakistan.     

Recent trends using natural polymeric nanofibers as supports for enzyme immobilization and catalysis

All the disciplines of science, especially biotechnology, have given continuous attention to the area of enzyme immobilization. However, the structural support made by material science intervention determines the performance of immobilized enzymes. Studies have proven that nanostructured supports can maintain better catalytic performance and improve immobilization efficiency. The recent trends in the application of nanofibers using natural polymers for enzyme immobilization have been addressed in this review article. A comprehensive survey about the immobilization strategies and their characteristics are highlighted. The natural polymers, e.g., chitin, chitosan, silk fibroin, gelatin, cellulose, and their blends with other synthetic polymers capable of immobilizing enzymes in their 1D nanofibrous form, are discussed. The multiple applications of enzymes immobilized on nanofibers in biocatalysis, biosensors, biofuels, antifouling, regenerative medicine, biomolecule degradation, etc.; some of these are discussed in this review article.     

Application of Microarray and Functional-Based Screening Methods for the Detection of Antimicrobial Resistance Genes in the Microbiomes of Healthy Humans

The aim of this study was to screen for the presence of antimicrobial resistance genes within the saliva and faecal microbiomes of healthy adult human volunteers from five European countries. Two non-culture based approaches were employed to obviate potential bias associated with difficult to culture members of the microbiota. In a gene target-based approach, a microarray was employed to screen for the presence of over 70 clinically important resistance genes in the saliva and faecal microbiomes. A total of 14 different resistance genes were detected encoding resistances to six antibiotic classes (aminoglycosides, β-lactams, macrolides, sulphonamides, tetracyclines and trimethoprim). The most commonly detected genes were erm(B), bla_TEM, and sul2. In a functional-based approach, DNA prepared from pooled saliva samples was cloned into Escherichia coli and screened for expression of resistance to ampicillin or sulphonamide, two of the most common resistances found by array. The functional ampicillin resistance screen recovered genes encoding components of a predicted AcrRAB efflux pump. In the functional sulphonamide resistance screen, folP genes were recovered encoding mutant dihydropteroate synthase, the target of sulphonamide action. The genes recovered from the functional screens were from the chromosomes of commensal species that are opportunistically pathogenic and capable of exchanging DNA with related pathogenic species. Genes identified by microarray were not recovered in the activity-based screen, indicating that these two methods can be complementary in facilitating the identification of a range of resistance mechanisms present within the human microbiome. It also provides further evidence of the diverse reservoir of resistance mechanisms present in bacterial populations in the human gut and saliva. In future the methods described in this study can be used to monitor changes in the resistome in response to antibiotic therapy.     

Recombination and population structure in Salmonella enterica

Salmonella enterica is a bacterial pathogen that causes enteric fever and gastroenteritis in humans and animals. Although its population structure was long described as clonal, based on high linkage disequilibrium between loci typed by enzyme electrophoresis, recent examination of gene sequences has revealed that recombination plays an important evolutionary role. We sequenced around 10% of the core genome of 114 isolates of enterica using a resequencing microarray. Application of two different analysis methods (Structure and ClonalFrame) to our genomic data allowed us to define five clear lineages within S. enterica subspecies enterica, one of which is five times older than the other four and two thirds of the age of the whole subspecies. We show that some of these lineages display more evidence of recombination than others. We also demonstrate that some level of sexual isolation exists between the lineages, so that recombination has occurred predominantly between members of the same lineage. This pattern of recombination is compatible with expectations from the previously described ecological structuring of the enterica population as well as mechanistic barriers to recombination observed in laboratory experiments. In spite of their relatively low level of genetic differentiation, these lineages might therefore represent incipient species.