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Objective: The Italian multicenter observational study depression in stroke (DESTRO) aimed to identify risk factors for poststroke depression (PSD) and quantify the likelihood of it arising in various categories of patients. Method: Mood evaluation was performed in 1064 consecutive stroke patients by means of Beck Depression Inventory and Visual Analog Mood Scale. Depressive symptoms were classified using the DSM-IV and revised WHO criteria for depression in the course of a neurological disorder. Results: Poststroke depression was seen in 36% of the survivors, with dysthymia by far the predominant form (80.7%). Female sex, disability, previous cerebrovascular or depressive episodes were significantly associated with an increased risk of depression. Combinations of these factors raised the risk of PSD exponentially, from 24.3 to 89.1%. The site of the stroke did not come into the uni- or multivariate analysis. Conclusion.: At admission, it is possible to predict the likelihood of PSD and quantify the relative risk.

We have synthesized by solution methods and characterized the lipopeptaibol metabolite LP237-F8 extracted from the fungus Tolypocladium geodes and five selected analogues with the Etn --> Aib or Etn --> Nva replacement at position 8 and/or a triple Gln --> Glu(OMe) replacement at positions 516, and 9 (Etn = Calpha-ethylnorvaline, Aib = alpha-aminoisobutyric acid, Nva = norvaline). Conformation analysis, performed by FT-IR absorption, NMR, and CD techniques, strongly supports the view that the six terminally blocked decapeptides are highly helical in solution. Helix topology and amphiphilic character are responsible for their remarkable membrane activity. At position 8 the combination of high hydrophobicity and Ca tetrasubstitution, as in the Etn-containing LP237-F8 metabolite, has a positive effect on membrane interaction.

Tributyltin (TBT), widely used as an antifouling biocide, is the most abundant pesticide in coastal environments. One of its main toxic effects is immunosuppression in both vertebrates and invertebrates. At sublethal doses of TBT, phagocytes lose their ability to move towards and ingest foreign particles. For short-term cultures of haemocytes (60 min at 25 ° C) of a marine invertebrate, the colonial ascidian Botryllus schlosseri, exposed to 10-5 M TBT, we previously reported dose- and time-dependent impairment of yeast phagocytosis and changes in cell morphology related to cytoskeleton disorganization. These effects are Ca2+-dependent, since inactivation of Ca2+-adenosine triphosphatase and a sustained increase in cytosolic Ca2+ occurred. As TBT can antagonize the effect of chlorpromazine, a specific calmodulin (CaM) inhibitor, and the co- presence of exogenous CaM and TBT in the incubation medium resulted in the absence of effects, we hypothesized an interaction between TBT and CaM. TBT may remove endogenous CaM from cell proteins, thus inactivating them and causing alteration of Ca2+ homeostasis. With the aim of confirming the hypothesis of a direct TBT-CaM interaction, we first studied the effects of co-incubation of TBT with other exogenous proteins on restoring the ability of phagocyte morphology. Although bovine serum albumin was never able to restore cell morphology, the effect of human spectrin was similar to that described for CaM, suggesting a common non-specific mechanism of action based on the interaction of TBT with exposed hydrophobic pouches. We also analysed the conformational changes of pure CaM (10-5 M) in the presence of various concentrations (10-4 to 10-3 M) of TBT and its degradation products, dibutyltin (DBT) and monobutyltin (MBT), by circular dichroism. Results indicate the dose- and time-dependent interaction of TBT with CaM. This interaction is a non-covalent interaction, probably hydrophobic in nature, between the aliphatic chains of TBT and the hydrophobic regions of Ca2+-activated CaM. DBT and MBT turned out to be less active in inducing CaM conformational changes, without any significant differences between the two compounds.

Bombolitins are five natural heptadecapeptides originally isolated from the venom of a bumblebee. They induce lysis of erythrocytes and liposomes and increase the activity of phospholipase A2 (PLA2), that plays an important role in the early steps of the inflammatory process. It has been proposed that PLA2 activation depends on the alteration of the physical state of the membrane. Bombolitin II folds into an alpha-helix in a membrane mimicking environment constituted by SDS micelles [Battistutta et al., Macromol. Chem. Phys., 196 (1995) 2827-2841]. In the present work, the topological orientation of the peptide relative to the micelle was determined, using three spin probes localized in different positions of the water/micelle system. The reduction in intensity of the 1H NMR signals clearly demonstrated that the peptide is located on the surface of the micelle, with its helical axis parallel to the interface. Only a small portion of the helix is exposed to the aqueous environment. Results from NMR experiments were confirmed by molecular dynamics simulations, performed using a two-phase water/decane simulation cell. The timescale for the reorientation of the peptide was between 120 and 450 ps, depending on the starting position of the peptide.

It was recently shown that the covalent tethering of the N-terminus of parathyroid hormone (PTH )to the seventh helical bundle of the G-protein coupled PTH-receptor (PTH1R)leads to autoactivation [Shimizu et al., J.Biol.Chem. 275 (2000) 19456–19460]. Here, we have developed molecular models for the interaction of PTH(1–11)tethered to PTH1R and refined them with molecular dynamics simulations. The starting structure of the ligand/receptor complex is based on experimental data from a series of spectroscopic structural studies of PTH(1–34)and the extracellular domains of PTH1R and intermolecular contact points derived from photoaffinity labeling. The resulting PTH1R/[Arg11]PTH(1–11)complex has the N-terminus of PTH interacting with residues of the third extracellular loop of PTH1R, as a possible mode for receptor activation. The hydrophobic residues leucine-5 and methionine-8,centrally located in the N-terminal alpha-helix of PTH(1–11), are located in deep, well-defined hydrophobic pockets in the central core of the seventh helical bundle, consistent with the requirement of these amino acids for autoactivation. We postulate that the improved signaling properties of [Arg]PTH(1–11)over wild type PTH(1–11)is due to a stable hydrogen bond between Arg11 and E444, at the beginning of TM7. The model provides atomic insight into currently available biochemical data as well as numerous putative ligand/receptor interactions, and thereby may further the rational design of reduced-size PTH agonists at the PTH1 receptor.

Bovine beta-lactoglobulin represents an interesting example of context-dependent secondary structure induction. In fact, secondary structure predictions indicated that this beta-barrel protein has a surprisingly high alpha-helical preference, which was retained for short fragments. Cooperative transitions from the native beta-sheet to alpha-helical structures were additionally induced by organic solvents, in particular trifluoroethanol. As a result of this high alpha-helical preference, it has been proposed that non-native alpha-helical intermediates could be formed in the unfolding pathway of this protein. In order to provide a better understanding of the processes that underlie conformational plasticity in this protein, CD measurements in the presence of increasing amounts of urea and in the presence of organic solvents were performed. Urea unfolding studies, performed at pH 2.1 and 37 degrees C, revealed an apparent two-state transition, and afforded no evidence of non native alpha-helical intermediates. The protein treated with up to 6M urea, refolded to the native structure, while treatment with higher molar concentration urea, lead to partial misfolding. A 29-mer peptide covering the region of strands a and b of the intact protein, characterized by the presence of 4/3 heptad repeats, was synthesized and studied by CD in the presence of different solvents. On the basis of the obtained results, a mechanism was proposed to explain the structural transition from the beta to alpha structure, provoked by organic solvents in the intact protein.

The heptadecapeptides bombolitin I and bombolitin III are two members of a series of biologically active peptides postulated to be membrane active. In order to understand the effects of the membrane on the secondary structure of the peptides, we have carried out the conformational characterization of bombolitins I and III in the presence of SDS micelles using circular dichroism, nuclear magnetic resonance, and computer simulations. The characteristic bands in the circular dichroism spectra indicate an alpha-helix content of approximately 60% in bombolitin III and 70% in bombolitin I. The observation of NOE's quite distinctive for such secondary structure strongly supports the CD results. The conformational preferences of the two bombolitins derived from CD and NMR were then energetically refined with molecular dynamics simulations. The results from the spectroscopic examination were utilized as input for the simulations, the CD results for generation of the initial structure, and the NOE's as constraints during the simulations. The results from the different techniques employed are in complete agreement. Biochemistry

The heptadecapeptides bombolitin I and bombolitin III are two of a series of peptides postulated to be biologically active within a membrane environment. In the preceding paper [Bairaktari, E., Mierke, D.F., Mammi, S., & Peggion, E. (1990) Biochemistry (preceding paper in this issue)] the conformational preferences of these peptides in the presence of SDS surfactant micelles, a mimetic for biological membranes, were examined. During these studies the conformations of these peptides were investigated in aqueous solutions by circular dichroism and nuclear magnetic resonance. A large difference was observed for the two peptides. Bombolitin I lacks any observable secondary structure in aqueous solution, independent of temperature, pH, and concentration. In striking contrast, bombolitin III adopts a well-defined alpha-helix at concentrations greater than 1.3 mM. This is indeed surprising given the great similarity of the two peptides. The alpha-helix of bombolitin III is pH dependent, with a great decrease in the observed secondary structure at pH values below 3.5. This observation could only be due to the protonation of the Asp residue at the fifth position. These findings suggest that the secondary structure arises from molecular aggregation of bombolitin III through the formation of a salt bridge involving the Asp side chain. The alpha-helix observed at "high" concentration (greater than 2.5 mM) has been characterized by CD and by the NOE's measured throughout a majority of the peptide. The experimentally determined structure has been energy refined with restrained molecular dynamics. The conformational results from this study are then compared with the conformations found in the presence of surfactant micelles. Biochemistry