Spectroscopic Characterization of Two Soluble Transducers from the Archaeon Halobacterium salinarum

In the present study, structural aspects of the two soluble transducers, HtrX and HtrXI, from the archaeon H. salinarum have been examined using UV circular dichroism and steady-state fluorescence spectroscopies. Circular dichroism (CD) data indicate that both HtrX and HtrXI exhibit salt-dependent protein folding. Under low-ionic-strength conditions (0.2 M NaCl or KCl) the CD spectra of HtrXI is similar to that of the Gdn-HCl- or urea-denatured forms and is indicative of random coil structure. In contrast, the CD spectrum of HtrX under low-ionic-strength conditions contains roughly 85% -helical character, indicating a significant degree of folding. Addition of NaCl or KCl to solutions of HtrX or HtrXI results in CD features consistent with predominately -helical character (>95%) for both proteins. In addition, the transition points (i.e., ionic strengths at which the protein converts from random coil to -helical character) are quite distinct and dependent upon the type of salt present (i.e., either NaCl or KCl). Accessibility of tryptophan residues to the solvent was also examined for both HtrX and HtrXI in both folded and unfolded states using Kl quenching. The Stern–Volmer constants obtained suggest that the tryptophans (Trp35 in HtrX and both Trp47 and Trp74 in HtrXI) are partially exposed to the solvent, indicating that they are located near the surface of the protein in all three cases. Furthermore, fluorescence quenching with the single Trp mutants Trp74AIa and Trp47AIa of HtrXI indicates different environments for these two residues.     

Identification of protein C epitopes altered during its nanoencapsulation

Protein C is a plasmatic inhibitor which regulates the blood coagulation mechanism by modulating the anticoagulant response. The improvement of its bioavailability would be beneficial for the treatment of the disorders caused by its homozygous deficiency or by an other plasmatic inhibitor deficiency. In this context, the protein C encapsulation into biodegradable nanoparticles could be used to approach the problem. However, the method used to prepare the nanoparticles requires the use of ultrasonication and of an organic solvent such as methylene chloride which interferes with protein activity. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that neither ultrasonication nor methylene chloride, singly or in combination, led to protein C aggregation or cleavage. Thus, a binding study using an ELISA assay with four characterized monoclonal antibodies was carried out to identify the epitopes damaged by these experimental constraints. The correlation between the immunological assay and a functional one i.e. by the means of the assay of its anticoagulant activity (activated partial thromboplastin time) made it possible to show that protein C amino acids 166–169 of the activation peptide were probably altered after ultrasonication and methylene chloride treatment. Indeed, it is likely that these residues were no longer surface-exposed but had moved inside the protein core.     

Isolation and identification of a 92-kDa stress induced protein from Candida albicans

Mycopathologia - It was previously shown that the presence of estrogen enhances survival of Candida albicans under heat and oxidative stresses. A 92-kDa protein is inducible by heat shock and...     

A highly conserved nematode protein folding operon in Caenorhabditis elegans and Caenorhabditis briggsae

In the free-living model nematode, Caenorhabditis elegans, a protein-folding co-transcribed gene pair has previously been described. The degree and form of trans-splicing, orientation and spacing of the genes, and the co-ordinate co-expression of protein folding catalysts in the nematode's hypodermis indicated this to be a functionally important operon. This gene pair has now been cloned and compared in the related organism Caenorhabditis briggsae to identify evolutionarily conserved, functionally important features. The corresponding C. briggsae gene pair was found to share the operon-specific features, including sequence homology blocks in the upstream 5′ flanking regions. The intergenic regions were not conserved. The homology block closest to the translational initiation codon of the upstream gene was found to contain a known Ceanorhabbitis promoter element site, and may therefore be an important cis-regulatory region directing the hypodermis-specific expression of this operon gene of C. elegans. This study also provides further confirmation of the high degree of chromosomal synteny between these nematode species.     

Anopheles gambiae feeding and survival on honeydew and extra-floral nectar of peridomestic plants

It is widely believed that the malaria vector Anopheles gambiae Giles (Diptera: Culicidae) rarely or never feeds on sugar in nature. If so, the need for supplemental blood-feeding may be increased and this would help to explain why it is such an efficient malaria vector. Nonetheless, both sexes of this mosquito species readily imbibe and digest sugar solutions, and sugar is a staple of laboratory colonies. In this study, we investigated whether An. gambiae will feed on the extra-floral nectar of three common peridomestic plants in Africa, and on honeydew of the mealybug Pseudococcus longispinus (Targioni-Tozetti) (Hemiptera: Homoptera: Pseudococcidae), and how this affects survivorship. We found that both males and females of An. gambiae provided with vegetative parts of cassava (Manihot esculenta Crantz) survived as well (x = 26.3 and 19.2 days, respectively) as they did on 50% sucrose solution (x = 29.7 and 24.3 days, respectively) and much longer than they did on water alone (x = 1.8 days, both sexes). Females provided with mealybug honeydew also lived substantially longer (x = 16.5 days) than those on water alone. Males and females provided with vegetative parts of castorbean (Ricinus communis L.) also survived much longer (x = 12.7 and 7.8 days, respectively) than on water, but those provided with flowering lantana (Lantana camara L.) did not. Anthrone tests of females after one night of exposure to these potential energy sources confirmed that they obtained fructose from cassava, from mealybug honeydew, and from non-flowering castorbean, but not from lantana or from castorbean lacking its petiolar nectaries. Previous laboratory studies had shown that sugar availability affects the survival and biting frequency of An. gambiae. It now appears that this mosquito can locate natural sources of plant sugar readily and utilize them effectively. Nectar-producing plants in the domestic environment may play a significant role in this mosquito's energy budget and malaria vectorial capacity.     

Species-specific variation in the importance of the spectral quality gradient in canopies as a signal for photosynthetic resource partitioning

BACKGROUND AND AIMS: Plants adjust the distribution of photosynthetic capacity and chlorophyll to canopy density. The importance of the gradient in the red : far-red ratio (R : FR) relative to the irradiance gradient was studied for its perception with respect to this partitioning of photosynthetic resources. Whether the relative importance of these two signals varied between six species of different growth habit (Phaseolus vulgaris, Lysimachia vulgaris, Hedera helix, Ficus benjamina, Carex acutiformis and Brachypodium pinnatum) was investigated further. METHODS: Single leaves of plants were shaded in daylight by a spectrally neutral filter or a leaf. In another experiment, leaves were treated with supplemental FR. In most cases, treatment effects were evaluated after 2 weeks. KEY RESULTS: Nitrogen and photosynthetic capacity (Amax) per leaf area, parameters pertaining to between-leaf resource partitioning, were strongly reduced in neutral shade but not additionally by spectral leaf shade. Supplemental FR reduced these parameters also, except in Carex. Acceleration of induction of senescence was observed in spectral leaf shade in primary bean leaves. Amax per unit chlorophyll, a parameter pertaining to within-leaf resource partitioning, was reduced in neutral shade, but not in spectral leaf shade or supplemental FR. CONCLUSIONS: Signalling mechanisms associated with perception of the R : FR gradient in canopies were less important than those associated with the irradiance gradient for between-leaf and within-leaf partitioning of photosynthetic resources. The relative importance of the signals differed between species because Carex was the only species for which no indications were found for an involvement of the spectral gradient in perception of canopy density.     

Role of protein in the primary step of the photoreaction of yellow protein

We show the unexpectedly important role of the protein environment in the primary step of the photoreaction of the yellow protein after light illumination. The driving force of the trans-to-cis isomerization reaction was analyzed by a computational method. The force was separated into two different components: the term due to the protein-chromophore interaction and the intrinsic term of the chromophore itself. As a result, we found that the contribution from the interaction term was much greater than that coming from the intrinsic term. This accounts for the efficiency of the isomerization reaction in the protein environment in contrast to that in solution environments. We then analyzed the relaxation process of the chromophore on the excited-state energy surface and compared the process in the protein environment and that in a vacuum. Based on this analysis, we found that the bond-selectivity of the isomerization reaction also comes from the interaction between the chromophore and the protein environment.     

Analysis of 6-hydroxy-2-aminocaproic acid (HACA) as a specific marker of protein oxidation: The use of <Emphasis Type="Italic">N(O,S)</Emphasis>-ethoxycarbonyl trifluoroethyl ester derivatives and gas chromatography/mass spectrometry

Summary. An alteration of low density lipoprotein (LDL) apolipoprotein (apo) B-100 structure by direct oxidative modification is an important mechanism involved in atherogenesis. There is difficulty in quantifying this type of modification because a lack of specific assays. The use of N(O,S)-ethoxycarbonyl trifluoroethyl amino acid esters for a rapid and sensitive determination of 6-hydroxy-2-aminocaproic acid (HACA), a highly specific marker of metal catalyzed protein oxidation, by using standard gas chromatography/electron impact mass spectrometry, is discussed. The derivatives are formed by the unlabored reaction of amino acids with ethylchloroformate plus trifluoroethanol plus pyridine. Femtomole levels of HACA can be reproducible measured in different LDL preparations subjected to oxidative damage in the presence of iron or copper. HACA determination compares well with the measurement of carbonyl groups that are generally accepted as a nonspecific index of protein oxidation. Thus, the method could prove to be a sensitive assay for studying specific apoB-100 modification.     

Proteins interacting with the tuberous sclerosis gene products

Summary. Tuberous sclerosis (TSC) is an autosomal dominant tumor suppressor gene syndrome affecting about 1 in 6000 to 10000 individuals. The genes, TSC1, encoding hamartin, and TSC2, encoding tuberin are responsible for TSC. Since their identification 1997 and 1993 respectively, a variety of different functions have been described for the TSC gene products. Hamartin and tuberin form a complex, providing a tentative explanation for the similar disease phenotype in TSC patients with mutations in either of these genes. In addition, associations of hamartin or tuberin with several different proteins have been demonstrated. In this review, we summarize the current knowledge on hamartin- and tuberin-interacting proteins and discuss their role for the understanding of the functions of the TSC gene products.     

Biochemical mechanisms for a possible involvement of the transglutaminase activity in the pathogenesis of the polyglutamine diseases: Minireview article

Summary. Transglutaminases are a family of enzymes which show the common capacity to catalyse the cross-linking of protein substrates. Some members of this family of enzymes are also capable to catalyse other chemical reactions for the cell life. The distribution and the role of these enzymes have been studied in numerous cell types and tissues, but only recently their expression and functions started to be investigated in the Nervous System. One of the main biochemical properties of the Transglutaminase enzymes is to form large protein aggregates that are insoluble in all known protein detergents. Recently, the Transglutaminase activity has been hypothesised to be involved in the pathogenetic mechanisms responsible for the formation of cellular inclusions present in the Corea Major and in other polyglutamine diseases. In this review we describe the biochemical mechanisms by which the Transglutaminases could play a critical role in the physiopathology of the polyglutamine diseases.