Conformational studies of a benzodiazepine-like peptide in SDS micelles by circular dichroism, 1H NMR and molecular dynamics simulation

The conformation of a benzodiazepine-like decapeptide corresponding tothe YLGYLEQLLR fragment of a casein has been examined in a sodium dodecylsulfate micellar medium using circular dichroism, two-dimensional¹H NMR spectroscopy and restrained molecular dynamicssimulation. The decapeptide adopts an amphipathic 3₁₀-helicoidstructure in which theE⁶···R¹⁰ ionic bridgestabilizes the C-terminus.     

Sperm motility in turbot,Scophthalmus marimus: initiation of movement and changes with time of swimming characteristics

Synopsis Turbot sperm motility is observed using dark field microscopy and stroboscopic illumination combined with video recording. Sperm motility is triggered by dilution of spermatozoa in sea water or in non ionic media (glucose or saccharose), presenting osmotic pressure ranging from 300 to 2100 mOsmol. The percentage of motile spermatozoa reaches 100% under conditions of osmotic pressure of 300 to 1100 mOsmol and pH close to 8.0. In full sea water, glucose or saccharose solutions an agglutination of spermatozoa is observed; this is prevented by addition of bovine serum albumin (5 mg ml^–1). Immediately after transfer in activation solutions, 100% spermatozoa are motile in most samples freshly stripped. This percentage drops suddenly between 15 and 30% after 70 to 100 sec. The beat frequency remains at a constant value of 50 Hz during 40 s post activation and then drops suddenly between 15 and 30 Hz. The spermatozoa velocity is about 200 micrometers s^–1 during 30 to 40 s and then declines to a stable value of 100 micrometers s^–1 at 50 s post activation. After 1.20 mn, more and more spermatozoa become motionless. The minimum calculated and averaged distance covered during 1.20 min, is about 12 mm. The high performances of turbot spermatozoa motility are interpreted as a compensatory mechanism for the low sperm production.     

Multiple modes of ligand recognition: Crystal structures of cyclin-dependent protein kinase 2 in complex with ATP and two inhibitors,olomoucine and isopentenyladenine

Cyclin-dependent kinases (CDKs) are conserved regulators of the eukaryotic cell cycle with different isoforms controlling specific phases of the cell cycle. Mitogenic or growth inhibitory signals are mediated, respectively, by activation or inhibition of CDKs which phosphorylate proteins associated with the cell cycle. The central role of CDKs in cell cycle regulation makes them a potential new target for inhibitory molecules with anti-proliferative and/or anti-neoplastic effects. We describe the crystal structures of the complexes of CDK2 with a weakly specific CDK inhibitor, N6-(δ2-isopentenyl)adenine, and a strongly specific inhibitor, olomoucine. Both inhibitors are adenine derivatives and bind in the adenine binding pocket of CDK2, but in an unexpected and different orientation from the adenine of the authentic ligand ATP. The N6-benzyl substituent in olomoucine binds outside the conserved binding pocket and is most likely responsible for its specificity. The structural information from the CDK2-olomoucine complex will be useful in directing the search for the next generation inhibitors with improved properties. © 1995 Wiley-Liss, Inc.     

pAMβ1 resolvase has an atypical recombination site and requires a histone-like protein HU

The broad-host-range plasmid pAMβ1 from Gram-positive bacteria encodes a resolvase, designated Resβ, which shares homology with the proteins of the resolvase—invertase family. Here we report the purification and in vitro characterization of Resβ. This resolvase is particular in two aspects: it has an atypical binding site and requires a cofactor to promote resolution in vitro . Resβ binds to two regions within its resolution site res . One contains two inverted repeats (R1 and R2), the other contains only one repeat (R3). The cofactor required for resolution in vitro is present in crude extracts of both Bacillus subtilis and Escherichia coli and can be substituted by the E. coli histone-like protein HU. The possible mode of action of HU in the resolution process is discussed.     

A three-step purification of human alpha 1-acid glycoprotein

alpha 1-Acid glycoprotein (AGP) was purified to homogeneity by a 3-step procedure using pseudo-ligand affinity chromatography on immobilized Cibacron blue F3GA, Procion red HE3B, and preparative column isoelectric focusing. The overall yield of the combined techniques was 88%. Analysis of the purified AGP by lectin affinity chromatography on immobilized Con A and immunoaffino-electrophoresis indicated that the most acidic form did not interact with the lectin, while the two more basic fractions possessed different affinities for Con A. In addition, 3 different populations of AGP were clearly separated by Con A affinity chromatography.     

Studies in vitro on the uptake and degradation of sodium hyaluronate in rat liver endothelial cells.

Rat liver endothelial cells in primary cultures at 7 degrees C bind radioactively labelled sodium hyaluronate (HA; Mr 400 000) specifically and with high affinity (Kd = 6 X 10(-11) M). Maximal binding capacity is approx. 10(4) molecules per cell. Inhibition experiments with unlabelled HA and oligosaccharides from HA indicate that each molecule is bound by several receptors acting co-operatively and that the single receptor recognizes a tetra- or hexa-saccharide sequence of the polysaccharide. At 37 degrees C the liver endothelial cells endocytose the HA. The process combines the features of a receptor-mediated and a fluid-phase endocytosis. The rate of internalization does not show any saturation with increasing HA concentration, but is approximately proportional to the polysaccharide concentration at and above the physiological concentration. At 50 micrograms of free HA/l each liver endothelial cell accumulates 0.1 fg of the polysaccharide/min. Fluorescent HA accumulates in perinuclear granules, presumably lysosomes. Degradation products from HA appear in the medium about 30 min after addition of the polysaccharide to the cultures. The radioactivity from HA containing N-[3H]acetyl groups or 14C in the sugar rings is recovered mainly as [3H]acetate and [14C]acetate respectively. Estimations of the capacity of liver endothelial cells to internalize and degrade HA in vitro indicate that these cells may be primarily responsible for the clearance of HA from human blood in vivo.     

A functional evenness index for microbial ecology

Microbial ecologists attempting to describe community structures through the use of synthetic parameters face enormous difficulties. These stem in part from the necessity of using standard taxonomic reference levels in a field where the species level is poorly defined. This paper presents an attempt to obviate this problem. A “functional evenness” index (E) is defined using information measures; it is based directly on the characteristics of the bacteria, as determined, for example, with the API 20B method. Comparisons of this index with classic structure indices, such as taxonomic evenness (Pielou) or systematic dominance (Hulburt), show that it behaves like an evenness index, while bypassing the taxonomic study required before computation of the classic indices. Its use is illustrated with samples of aerobic heterotrophic bacteria obtained from brackish lagoon sediments. We are grateful to our statistician colleagues, Dr. Yves Lepage from Université de Montréal and Dr. André Plante from Université du Québec à Montréal. Both have contributed important elements to the discussion presented in this Appendix.