Differential irreversible insertion of protein kinase C into phospholipid vesicles by phorbol esters and related activators.

Incubation of protein kinase C (PKC) alpha with phorbol 12,13-dibutyrate and phospholipid vesicles promoted a time-dependent irreversible insertion of the enzyme into the vesicles and the generation of a calcium-independent kinase activity. Calcium neither caused insertion nor influenced the insertion induced by the phorbol ester. The effect was strongly dependent on the phosphatidylserine concentration in the vesicle and could also be supported by other anionic phospholipids. An analysis of the structure-activity relations of PKC activators for the calcium-independent kinase activity revealed marked relative differences in potencies for binding and for insertion. Compounds such as phorbol 13-myristate 12-acetate and mezerein were very efficient at inducing insertion. In contrast, 12-deoxyphorbol esters and diacylglycerol were relatively inefficient at inducing insertion, requiring higher concentrations than expected from their binding affinities. The insertion of PKC alpha depended substantially on the length of the aliphatic esters in the 12- and 13-positions of the phorbol derivatives, and once again, potencies for insertion and binding were not directly proportional. Our findings suggest two different sites for ligand interaction on the molecule of PKC alpha with different structure-activity requirements. We speculate that the differential ability of compounds to promote insertion could contribute to the documented marked differences in the biological behavior of PKC activators.     

Spatiotemporal testing and modeling of catfish retinal neurons.

The responses of retinal neurons depend on the interaction of both temporal and spatial aspects of a light stimulus. We developed a linear spatiotemporal model of receptor and horizontal cell layers in the catfish retina based on reciprocal interactions between both layers and coupling within each. Horizontal cell transfer properties were measured experimentally using white-noise intensity modulated light spots of different diameters and were compared with analytical predictions based on the model. Good agreement was obtained with a reasonable choice of model space-constants and feedback parameters. Furthermore, the same set of parameter values determined from spot experiments enabled accurate prediction of experimental horizontal cell responses to traveling gratings. The proposed feedback connections from horizontal cells to receptors quicken the time-course of responses in both layers and sharpen receptive fields.     

Glycosylation reaction of unprotected sugars with hydroxyalkylthymine

Under mild conditions (Lewis acid/solvent/room temperature), the reaction of unprotected glucose, deoxyribose or xylose with hydroxylalkylthymine gives selectively nucleoside analogs with a spacer arm between sugar and base moiety. Experimental conditions (Lewis acid, solvent) for this new strategy leading to nucleoside analogs synthesis are discussed.     

D1 protein variants in Photosystem II from Thermosynechococcus elongatus studied by low temperature optical spectroscopy

In Photosystem II (PSII) from Thermosynechococcus elongatus, high-light intensity growth conditions induce the preferential expression of the psbA₃ gene over the psbA₁ gene. These genes encode for the D1 protein variants labeled D1:3 and D1:1, respectively. We have compared steady state absorption and photo-induced difference spectra at < 10 K of PSII containing either D1:1 or D1:3. The following differences were observed. (i) The pheophytin Q_x band was red-shifted in D1:3 (547.3 nm) compared to D1:1 (544.3 nm). (ii) The electrochromism on the Pheo_D1 Q_x band induced by Q_A⁻ (the C550 shift) was more asymmetric in D1:3. (iii) The two variants differed in their responses to excitation with far red (704 nm) light. When green light was used there was little difference between the two variants. With far red light the stable (t_1/2 > 50 ms) Q_A⁻ yield was ∼ 95% in D1:3, and ∼ 60% in D1:1, relative to green light excitation. (iv) For the D1:1 variant, the quantum efficiency of photo-induced oxidation of side-pathway donors was lower. These effects can be correlated with amino acid changes between the two D1 variants. The effects on the pheophytin Q_x band can be attributed to the hydrogen bond from Glu130 in D1:3 to the 13¹-keto of Pheo_D1, which is absent for Gln130 in D1:1. The reduced yield with red light in the D1:1 variant could be associated with either the Glu130Gln change, and/or the four changes near the binding site of P_D1, in particular Ser153Ala. Photo-induced Q_A⁻ formation with far red light is assigned to the direct optical excitation of a weakly absorbing charge transfer state of the reaction centre. We suggest that this state is blue-shifted in the D1:1 variant. A reduced efficiency for the oxidation of side-pathway donors in the D1:1 variant could be explained by a variation in the location and/or redox potential of P⁺.     

Oxygen-evolving Photosystem II core complexes: a new paradigm based on the spectral identification of the charge-separating state, the primary acceptor and assignment of low-temperature fluorescence.

We review our recent low-temperature absorption, circular dichroism (CD), magnetic CD (MCD), fluorescence and laser-selective measurements of oxygen-evolving Photosystem II (PSII) core complexes and their constituent CP 4 3, CP 47 and D1/D2/cytb(559) sub-assemblies. Quantitative comparisons reveal that neither absorption nor fluorescence spectra of core complexes are simple additive combinations of the spectra of the sub-assemblies. The absorption spectrum of the D1/D2/cytb(559) component embedded within the core complex appears significantly better structured and red-shifted compared to that of the isolated sub-assembly. A characteristic MCD reduction or 'deficit' is a useful signature for the central chlorins in the reaction centre. We note a congruence of the MCD deficit spectra of the isolated D1/D2/cytb(559) sub-assemblies to their laser-induced transient bleaches associated with P 680. A comparison of spectra of core complexes prepared from different organisms helps distinguish features due to inner light-harvesting assemblies and the central reaction-centre chlorins. Electrochromic spectral shifts in core complexes that occur following low-temperature illumination of active core complexes arise from efficient charge separation and subsequent plastoquinone anion (Q(A)(-)) formation. Such measurements allow determinations of both charge-separation efficiencies and spectral characteristics of the primary acceptor, Pheo(D1). Efficient charge separation occurs with excitation wavelengths as long as 700 nm despite the illuminations being performed at 1.7 K and with an extremely low level of incident power density. A weak, homogeneously broadened, charge-separating state of PSII lies obscured beneath the CP 47 state centered at 690 nm. We present new data in the 690-760 nm region, clearly identifying a band extending to 730 nm. Active core complexes show remarkably strong persistent spectral hole-burning activity in spectral regions attributable to CP 43 and CP 47. Measurements of homogeneous hole-widths have established that, at low temperatures, excitation transfer from these inner light-harvesting assemblies to the reaction centre occurs with approximately 70-270 ps(-1) rates, when the quinone acceptor is reduced. The rate is slower for lower-energy sub-populations of an inhomogeneously broadened antenna (trap) pigment. The complex low-temperature fluorescence behaviour seen in PSII is explicable in terms of slow excitation transfer from traps to the weak low-energy charge-separating state and transfer to the more intense reaction-centre excitations near 685 nm. The nature and origin of the charge-separating state in oxygen-evolving PSII preparations is briefly discussed.     

Hydrophobic plastics films synthesis by mean of agaroid acylation with lauroyl chloride in the N,N-dimethylacetamide homogeneous system

The synthesis of hydrophobic plastic films was performed by acylation of agaroids with lauroyl chloride in the N,N-dimethylacetamide homogeneous system. All the plastic films were characterized by FT-IR spectroscopy and their degrees of substitution (DS) was deduced from their ¹H-NMR spectra. In addition, thermomechanical feature of plastic films were analyzed and compared to those obtained from other kinds of hydrophobic plastic films. Latin square design of experiments helped us to determine optimized experimental conditions and identify the most important factors. Hence, mild conditions of acylation (90∘C, 1 eq/OH of lauroyl chloride, 1 eq/OH of 4-dimethylaminopyridine, 5,min) led to the production of highly substituted plastic films (DS = 3.62; maximum 4) with a high weight yield (211%) displaying mechanical properties close to polyethylene low density.     

Synthesis and properties of new bolaform and macrocyclic galactose-based surfactants obtained by olefin metathesis

A series of galactose-based surfactants with various structures likely to display new interesting properties were synthesized. Four monocatenary surfactants were elaborated by microwave-assisted galactosylation of undecanol or 10-undecenol. These compounds were slightly soluble in water. Their tensioactive properties were determined at 45 degrees C. Olefin metathesis was used to synthesize the two single-chain bolaforms from undec-10-enyl galactopyranosides; two pseudomacrocyclic bolaforms were prepared by grafting two carbamates at O-4 and O-4' sugar positions of the single-chain bolaforms. These four surfactants are insoluble in water and undergo monolayer compression. Cyclization of these bolaforms by olefin metathesis led to macrocyclic surfactant analogues of archaeobacterial membrane components.     

Sperm Cryopreservation in Male Infertility Due to Genetic Disorders

Certain chromosomal and genetic anomalies, such as Klinefelter syndrome (47,XXY) and Y chromosome microdeletions, have been reported as potential causes of a progressive impairment of spermatogenesis. In these cases cryoconservation of ejaculated or testicular sperm represent a valuable tool for the preservation of fertility. However, dealing with genetic disorders, the transmission of genetic anomalies has to be taken into consideration. It is therefore important to be aware about the clinical importance and the related genetic risks of these anomalies. In this article we describe the clinical significance of these diseases.