Interconvertible geometric isomers of Plasmodium falciparum dihydroorotate dehydrogenase inhibitors exhibit multiple binding modes

Two new tricyclic β-aminoacrylate derivatives (2e and 3e) have been found to be inhibitors of Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) with Ki 0.037 and 0.15 μM respectively. ¹H and ¹³C NMR spectroscopic data show that these compounds undergo ready cis-trans isomerisation at room temperature in polar solvents. In silico docking studies indicate that for both molecules there is neither conformation nor double bond configuration which bind preferentially to PfDHODH. This flexibility is favourable for inhibitors of this channel that require extensive positioning to reach their binding site.     

Affinity and specificity requirements for the first Src homology 3 domain of the Crk proteins.

The specificity of SH3 domain complex formation plays an important role in determining signal transduction events. We have previously identified a highly specific interaction between the first CrkSH3 domain [CrkSH3(1)] and proline-rich sequences in the guanine nucleotide exchange factor C3G. A 10 amino acid peptide derived from the first proline-rich sequence (P3P4P5A6L7P8P9K10K11R12) bound with a Kd of 1.89 +/- 0.06 microM and fully retained the high affinity and unique selectivity for the CrkSH3(1) domain. Mutational analysis showed that P5, P8, L7 and K10 are critical for high affinity binding. A conservative mutation, K10R, significantly decreased the affinity for the CrkSH3(1) domain while increasing the affinity for Grb2. Comparative binding studies with the K10R and K10A mutant peptides to c-Crk and v-Crk further suggested that K10 binds via a charge-dependent and a charge-independent interaction to the RT loop of the CrkSH3(1) domain. Besides determining important structural features necessary for high affinity and specificity binding to the CrkSH3(1) domain, our results also demonstrate that a conservative mutation in a single amino acid can significantly alter the specificity of an SH3 binding peptide.     

Influence of exogenous steroids, nutrition and calf removal on reproductive performance of anestrous beef cows

Methods for inducing a fertile estrus in anestrous beef cows suckling calves were explored in three studies. Trial 1 . Eighty-two primiparous and pluriparous Brahman-Hereford crossbred and Angus cows suckling calves were divided into four groups: 1) good body condition, high level of nutrition (GH); 2) good body condition, low level of nutrition (GL); 3) poor body condition, high level of nutrition (PH); 4) poor body condition, low level of nutrition (PL). All cows received the Shang Treatment (S). Following treatment, more cows (P<0.05) ovulated in the GH group (95%) than either the GL (61%), PH (58%), or PL (53%) group. Pregnancy rates were low and were not different between groups following the insemination taking place 54 hours after implant removal or after cows had been bred for 21 days. Trial 2 . One hundred fourteen two-year-old Santa Gertrudis-Hereford crossbred cows in poor body condition were divided into four groups. Forty-nine cows received S treatment and 65 cows served as controls (C) with approximately half of each group fed a high (H) or a medium (M) level of nutrition (SH, SM, CH, CM). One cow in the SH group ovulated and was pregnant 21 days after breeding. The remaining cows failed to exhibit estrus, ovulate or become pregnant. Trial 3 . Cows used in this trial were those from Trial 2 which did not exhibit estrus nor had a palpable corpus luteum present 21 days after implant removal. Cows were divided into 1) control (C), 2) wean (W), 3) wean + Syncro-Mate-B (W+SMB) and 4) Syncro-Mate-B + wean (SMB+W). The percentage of cows ovulating by 2 and 40 days after the start of breeding was similar (P>0.05) in the W, W+SMB, and SMB+W groups but all were greater (P<0.05) than the percentage of cows ovulating in the C groups. Pregnancy rates of cows in the W+SMB and SMB+W groups were higher (P<0.05) than pregnancy rates of cows in the W and C groups by two days after the start of breeding. Pregnancy rates 40 days after the start of breeding were higher (P<0.05) for cows in the W, W+SMB) and SMB+W groups than for cows in the C group.     

An histological and immunocytochemical study of the neuroendocrine cells in the intestine of Podarcis hispanica Steindachner, 1870 (Lacertidae)

Summary Numerous endocrine cells can be observed in the gut of the lizard Podarcis hispanica after application of the Grimelius silver nitrate technique. The argyrophilic endocrine cells are usually tall and thin in the small intestine but short, basal, and round in the large intestine. Eleven types of immunoreactive endocrine cells have been identified by immunocytochemical methods. Numerous serotonin-, caerulein/gastrin/cholecystokinin octapeptide-and peptide tyrosine-tyrosine-immunoreactive cells; a moderate number of pancreatic polypeptide-, neurotensin-, somatostatin-, glucagon-like peptide-1-and glucagon-immunoreactive cells, and few cholecystokinin N-terminal-and bombesin-immunoreactive cells were found in the epithelium of the small intestine. Coexistence of glucagon with GLP-1 or PP/PYY has been observed in some cells. In the large intestine a small number of serotonin-, peptide tyrosine-tyrosine-, pancreatic polypeptide-, neurotensin-, somatostatin-and glucagon-like peptide-1-immunoreactive cells were detected. Vasoactive intestinal peptide immunoreactivity was found in nerve fibers of the muscular layer. Substance P-immunoreactive nerve fibers were detected in lamina propria, submucosa and muscular layer. Chromogranin A-immunoreactive cells were observed throughout the intestine, although in lower numbers than argyrophilic cells.     

Protein immobilization to alumina supports: II. Papain immobilization to alumina via organophosphate linkers

Particulate aluminum oxides (alumina) were examined as supports for the immobilization of the proteolytic enzyme papain. Two alumina supports termed C1 and CPC were derivatized using organic phosphate linkers to create free carboxyl groups using a two-step process. Papain binding to these derivatized aluminas was performed using the water soluble carbodiimide 1-ethyl-3-(dimethylaminopropyl) carbodiimide. Reactions were optimal at 10 mM carbodiimide. The immobilized protein showed similar kinetic constants when compared to the solution protein. The pH dependence and thermal stability were essentially identical. The immobilized papain showed a blue shift in the intrinsic fluorescence emission maxima. Papain modified with the active site-specific fluorescent probe acrylodan showed overlapping emission maxima. These results are interpreted as retention of the hydrophobic environment of the active site with a perturbation in the structure of the rest of the protein caused by its association with the negatively charged surface. (c) 1992 John Wiley & Sons, Inc.     

Improved Device for the Administration of Fungal Spores to Small Animals via the Respiratory Route

An improved device has been designed and constructed for the administration of fungal spores or other air-suspended particles to mice via inhalation. Fabricated from Lucite [0.25 inch (0.63 cm)], the device was built in the shape of a 6-inch (15.2-cm) cube with two 1-inch (2.5-cm) holes drilled in each side. Mice are restrained in plastic conical centrifuge tubes which fit into the holes by friction. The device is safe, easily cleaned and sterilized, and can accommodate up to eight animals at a single exposure.     

Genetic manipulation in cultivars of oilseed rape (Brassica napus) using Agrobacterium

Summary The response of oilseed rape cultivars to infection with Agrobacterium tumefaciens and A. rhizogenes and the possibility of regenerating genetically transformed oilseed rape plants were examined. The frequency at which Agrobacterium induced galls or hairy-roots on in vitro cultured plants ranged from 10% to 70%, depending on the cultivar. From galls induced by the tumorigenic strain T37, known to be strongly shoot inducing on tobacco, roots developed frequently. Occasionally, shoots formed and some of these produced tumour cell specific nopaline. Attempts to grow the transformed shoots into plants have so far been unsuccessful. Whole plants transformed with Ri-T-DNA, however, were regenerated. These had crinkled leaves and abundant, frequently branching roots that showed reduced geotropism, similar to previously isolated Ri T-DNA transformed tobacco and potato plants. The transformed oilseed rape plants flowered, but failed to form seeds.