Implications of adopting a biodiversity-based vulnerability index versus a shoreline environmental sensitivity index on management and policy planning along coastal areas.

In this study, a multi-criteria index was developed to assess anthropogenic stressors along the Mediterranean coastline. The index aimed at geo-locating pollution hotspots for informed decision making related to coastal zone management. The index was integrated in a Geographical Information System based geodatabase implemented at several pilot areas along the Northern (Italy and France), Eastern (Lebanon), and Southern (Tunisia) Mediterranean coastlines. The generated stressor maps were coupled with a biodiversity richness index and an environmental sensitivity index to produce vulnerability maps that can form the basis for prioritizing management and mitigation interventions towards the identification of pollution hotspots and the promotion of sustainable coastal zone management. The results identified significant differences between the two assessment methods, which can bias prioritization in decision making and policy planning depending on stakeholders' interests. The discrepancies emphasize the need for transparency and understanding of the underlying foundations behind vulnerability indices and mapping development.

Patterns of use of oral anticancer treatments in France: a Retrospective Analysis of Cancer Treatments given ORally from 2004 to 2012 (Re-ACTOR study).

BACKGROUNDS AND OBJECTIVES: Increasing use of oral anticancer treatments (OATs) in oncology is modifying the treatment paradigm for cancer. Nonetheless, available data on the pattern of use of OATs and its evolution over time are limited. The objective of this study was to describe the patterns of use of OATs in France from 2004 to 2012. METHODS: A retrospective analysis was performed using Oncology Analyzer, a physician survey database. All patients actively treated by an oral or an intravenous anticancer treatment between October 2004 and September 2012 were enrolled in the database. Descriptive analyses were performed by treatment category with a focus on the last year of collection and the evolution across the study period. RESULTS: From October 2011 to September 2012, a sample of 7426 patients treated by oral or intravenous active anticancer treatments was analyzed: 74% of patients receiving an OAT were diagnosed with a solid tumor, 52% of whom had a stage IV cancer. The use of OATs increased with age and was the highest in patients over 80 years. From 2004 to 2012, the proportion of cancer patients receiving OATs increased by four percentage points (from 28.4% to 32.5%). Additionally, for treatments available in both forms, a marked preference for oral formulations was observed. LIMITATIONS: The patterns and trend of use prior to 2004 were not addressed due to lack of information in the database. The use of a market research database is relevant for highly prevalent cancers but for rare cancers the sample size is limited, underlining the utility of using other data sources such as cancer registries. CONCLUSIONS: The Re-ACTOR study provides an overview of OAT use in France, which was prescribed to 32% of cancer patients in France in 2012, principally to older patients and to those with solid tumors and with metastatic disease.

Determinants of diarrhea prevalence in urban slums: a comparative assessment towards enhanced environmental management.

This study relies on a comparative assessment of diarrhea occurrence in two urban slums to identify salient factors influencing case prevalence. Primary data were collected from both areas using a structured closed-ended questionnaire coupled with bottled and public water quality sampling and analysis at households reporting diarrhea cases. The water quality analysis showed contamination at the household level due primarily to the location of water storage tanks, as well as in some brands of bottled water due to lack of enforcement of source monitoring. Descriptive statistics and chi-square distribution tests revealed significant difference in diarrhea cases in both study areas which was correlated with the educational level of household head, financial status, type of water storage tank, and corresponding cleaning frequency as well as the adoption of measures to treat water or the use of bottled water.

Public perception and economic implications of bottled water consumption in underprivileged urban areas.

This paper presents a comparative assessment of public perception of drinking water quality in two underprivileged urban areas in Lebanon and Jordan with nearly similar cultural and demographic characteristics. It compares the quality of bottled water to the quality of the drinking water supplied through the public network and examines the economic implications of bottled water consumption in the two study areas. Participants' perception of the quality of drinking water provided via the public network was generally negative, and bottled water was perceived to be of better quality in both areas, thus affecting drinking water preferences and consumption patterns. The results reveal that the quality of bottled water is questionable in areas that lack enforcement of water quality standards, thus adding to the burden of an already disadvantaged community. Both areas demonstrated a considerable cost incurred for purchasing bottled water in low income communities reaching up to 26 % of total income.

ADM1 performance using SS-OFMSW with non-acclimated inoculums.

This paper assesses the anaerobic digestion (AD) of the source-sorted organic fraction of municipal solid waste (SS-OFMSW). For this purpose, an experimental programme was implemented involving the operation and monitoring of two bench-scale anaerobic digesters, continuously fed with SS-OFMSW. The mathematical model (ADM1) was then applied to simulate the process of AD of SS-OFMSW. While start-up of the digesters was relatively slow, re-inoculation with cattle manure with effluent dilution reduced the acclimation period and achieved better stability, accommodating a feeding rate at an OLR = 2.39 kg TVS m(-3) day(-1). The high recorded methane gas production rate, reaching (0.1-2.5 m(3) CH(4)/m(3) reactor day), confirms the excellent biodegradability of the type of waste used (SS-OFMSW) and its suitability for AD. Satisfactory simulations of soluble chemical oxygen demand (COD), pH, and methane composition of biogas were obtained, whereas volatile fatty acid (VFA) concentrations in both reactors were over-predicted albeit capturing its general trend.

HIV-1 integrase and virus and cell DNAs: complex formation and perturbation by inhibitors of integration.

HIV-1 integrase (IN) catalyzes integration of viral DNA into cell DNA through 3'-processing of viral DNA and strand transfer reactions. To learn on binding of IN to DNAs and IN inhibition we applied spectroscopy (circular dichroism, fluorescence) in a simplified model consisting in a peptide analogue (K156) of alpha4 helix involved in recognition of viral and cell DNA; an oligonucleotide corresponding to the U5' LTR DNA end; and an inhibitor (TB11) of the diketo acid (DKA) family. Results extrapolated to IN show that: the enzyme binds viral DNA with high affinity and specificity, but cell DNA with low affinity and specificity; the affinity of TB11 for IN is high enough to impair the binding of IN to cell DNA, but not to viral DNA. This explains why TB11 is an inhibitor of strand transfer but not of 3'-processing. These results can help in the search of new IN inhibitors.

Identification of the substrate-binding exosites of two snake venom serine proteinases: molecular basis for the partition of two essential functions of thrombin.

The venom of the South American snake Bothrops jararaca contains two serine proteinases, bothrombin and the platelet-aggregating enzyme PA-BJ, which share 66% sequence identity. Each of these proteinases possesses one of the two essential procoagulant functions of thrombin-the clotting of fibrinogen and platelet aggregation. Thus, bothrombin clots fibrinogen but has no direct effect on platelets, unless in the presence of exogenous fibrinogen. PA-BJ induces platelet aggregation by interacting with the protease-activated platelet receptor without clotting fibrinogen. On the other hand, thrombin possesses two extended surfaces. One is composed of basic and hydrophobic residues (exosite I) and the other one of basic residues only (exosite II). These exosites are involved in the recognition of physiological macromolecular substrates. In order to identify the corresponding exosites in bothrombin and PA-BJ and understand the molecular basis of the partition of the two procoagulant functions of thrombin among the two snake venom enzymes, we used molecular modeling to obtain models of their complexes with their natural substrates fibrinogen and a fragment of the protease-activated platelet receptor, respectively. In analogy to thrombin, each of the enzymes presents two exosites. Nonetheless, the exosites contain a smaller proportion of basic residues than thrombin does (45-72%), reducing thus the functional diversity of the enzymes. In addition, the composition of exosite I is different in both enzymes. We identify those residues in exosite I that could contribute to the differences in specificity. Finally, allostery does not seem to mediate macromolecular substrate recognition by these enzymes.

Toxicity evolution of Vipera aspis aspis venom: identification and molecular modeling of a novel phospholipase A(2) heterodimer neurotoxin.

We report the simultaneous presence of two phospholipase A(2) (PLA(2)) neurotoxins in the venom of Vipera aspis aspis, the first such observation. One is monomeric and identical to ammodytoxin B of Vipera ammodytes ammodytes. Its presence may result from gene flux after interbreeding between V. aspis aspis and V. ammodytes ammodytes. The second, a novel heterodimer named vaspin, is very similar to vipoxin of Vipera ammodytes meridionalis and to PLA(2)-I of Vipera aspis zinnikeri. It may result from expression of preexisting genes, the acidic subunit evolving from an ancestor common to ammodytin I2 from V. ammodytes ammodytes, which we also found in V. aspis aspis.

Peptide inhibitors of HIV-1 integrase dissociate the enzyme oligomers.

Integration of HIV-1 genome into host cell chromosome is mediated by viral integrase (IN). The IN catalytic core (CC, IN(50-212)) dimerizes through mutual interactions of its alpha1 and alpha5 helices. Peptides INH1 and INH5 reproducing these helix sequences strongly inhibited IN. For instance, an IC(50) of 80 nM was determined for INH5 in integration assays using wild-type IN (wtIN). In size exclusion chromatography, INH1 and INH5 perturbed the association-dissociation equilibrium of both dmIN (IN(1-288)/F185K/C280S) and CC, leading to monomers as surviving species, while in circular dichroism, binding of peptides to dmIN altered the protein conformation. Thus, enzyme deactivation, subunit dissociation, and protein unfolding are events which parallel one another. The target of INH5 in the enzyme was then identified. In fluorescence spectroscopy, C(0.5) values of 168 and 44 nM were determined for the binding affinity of INH5 to IN and CC, respectively, at 115 nM subunit concentration, while interaction of INH5 with INH1 was found stronger than interaction of INH5 with itself (23 times larger in term of dissociation constants). These results strongly suggested that the alpha1 helix is the privileged target of INH5. The latter could serve as a lead for the development of new chemotherapeutic agents against HIV-1.

Identification and molecular structural prediction analysis of a toxicity determinant in the Bacillus sphaericus crystal larvicidal toxin.

The operon containing the genes encoding the subunits of the binary crystal toxin of Bacillus sphaericus strain LP1-G, BinA and BinB (41.9 kDa and 51.4 kDa, respectively), was cloned and sequenced. Purified crystals were not toxic to Culex pipiens larvae. Comparison of the amino-acid sequences of this strain (Bin4) with those of the three other known toxin types (Bin1, Bin2 and Bin3) revealed mutations at six positions, including a serine at position 93 of BinA4, whereas all other types of BinA toxin from B. sphaericus had a leucine at this position. Reciprocal site-directed mutagenesis was performed to replace this serine in BinA4 from LP1-G with a leucine and the leucine in the BinA2 protein from strain 1593 with a serine. Native and mutated genes were cloned and overexpressed. Inclusion bodies were tested on C. pipiens larvae. Unlike the native Bin4 toxin, the mutated protein was toxic, and the reciprocal mutation in Bin2 led to a significant loss of toxicity. In vitro receptor-binding studies showed similar binding behaviour for native and mutated toxins. In the absence of any experimental data on the 3D structure of these proteins, sequence analysis and secondary-structure predictions were performed. Amino acid 93 of the BinA polypeptide probably belongs to an alpha helix that is sensitive to amino-acid modifications. Position 93 may be a key element in the formation of the BinA-BinB complex responsible for the toxicity and stability of B. sphaericus Bin toxins.

Self-association of an amphipathic helix peptide inhibitor of HIV-1 integrase assessed by electro spray ionization mass spectrometry in trifluoroethanol/water mixtures.

Establishing the auto-associative properties of a molecule in solution can be important for determination of its structure and function. EAA26 (VESMNEELKKIIAQVRAQAEHLKTAY) has been designed to inhibit HIV-1 integrase via formation of a stable coiled-coil structure with a nearly homologous segment in the enzyme. The latter catalyzes the permanent incorporation of a DNA copy of the retrovirus genome into host cell DNA, and is thus essential to the life of the retrovirus. This makes integrase an obvious drug target in the therapy of AIDS. The present work has demonstrated, using electrospray ionization mass spectrometry (ESI-MS), that EAA26 is monomeric in pure water, and tetrameric and dimeric at respectively low and medium concentrations of 2,2,2-trifluoroethanol (TFE), and again monomeric at higher TFE concentrations. Thus, the apolar solvent TFE may contribute to either stabilization or disruption of the intermolecular hydrophobic contacts depending on its concentration in aqueous solution. Previous NMR and ultracentifugation results are thus confirmed, indicating the reliability of ESI-MS for defining the self-association state of biologically relevant peptides in both water and organic-water solutions.

Molecular basis for the partition of the essential functions of thrombin among snake venom serine proteinases: the case of thrombin-like enzymes.

Thrombin is a mammalian serine proteinase that plays a prominent role in the maintenance and regulation of hemostasis through its interaction with various substrates and/or ligands. The venoms of several snakes contain glycosylated serine proteinases that have been recognized to possess one or more of the essential activities of thrombin on fibrinogen (Fg) and/or platelets. These proteinases share about 60% sequence identity. One class of snake venom serine proteinases are those known as thrombin-like (TLE), named after their ability to directly clot Fg in order to preferentially produce fibrinopeptide A, fibrinopeptide B or both. To understand the molecular basis of this phenomenon, the corresponding amino acid sequences and molecular structures need to be analyzed. Given the absence of experimentally determined tertiary structures of snake venom, TLEs, three-dimensional molecular models should prove useful in this context. Towards this goal, we obtained models of snake venom TLEs that used TSV-PA as template, TSV-PA being the only snake venom serine proteinase whose crystal structure is known to date. Along with a comparative sequence analysis the models contribute to the identification and description of thrombin-homologous or alternative binding sites, helping thus to understand differences in specificity.

Combining phage display and molecular modeling to map the epitope of a neutralizing antitoxin antibody.

Crotoxin is a potent presynaptic neurotoxin from the venom of the rattlesnake Crotalus durissus terrificus. It is composed of the noncovalent and synergistic association of a weakly toxic phospholipase A2, CB, and a nontoxic three-chain subunit, CA, which increases the lethal potency of CB. The A-56.36 mAb is able to dissociate the crotoxin complex by binding to the CA subunit, thereby neutralizing its toxicity. Because A-56.36 and CB show sequence homology and both compete for binding to CA, we postulated that A-56.36 and CB had overlapping binding sites on CA. By screening random phage-displayed libraries with the mAb, phagotopes bearing the (D/S)GY(A/G) or AAXI consensus motifs were selected. They all bound A-56.36 in ELISA and competed with CA for mAb binding, although with different reactivities. When mice were immunized with the selected clones, polyclonal sera reacting with CA were induced. Interestingly, the raised antibodies retained the crotoxin-dissociating effect of A-56.36, suggesting that the selected peptides may be used to produce neutralizing antibodies. By combining these data with the molecular modeling of CA, it appeared that the functional epitope of A-56.36 on CA was conformational, one subregion being discontinuous and corresponding to the first family of peptides, the other subregion being continuous and composed of amino acids of the second family. Phage-displayed peptides corresponding to fragments of the two identified regions on CA reacted with A-56.36 and with CB. Our data support the hypothesis that A-56.36 and CB interact with common regions of CA, and highlight residues which are likely to be critical for CA-CB complex formation.

The contribution of residues 192 and 193 to the specificity of snake venom serine proteinases.

Snake venom serine proteinases, which belong to the subfamily of trypsin-like serine proteinases, exhibit a high degree of sequence identity (60-66%). Their stringent macromolecular substrate specificity contrasts with that of the less specific enzyme trypsin. One of them, the plasminogen activator from Trimeresurus stejnegeri venom (TSV-PA), which shares 63% sequence identity with batroxobin, a fibrinogen clotting enzyme from Bothrops atrox venom, specifically activates plasminogen to plasmin like tissue-type plasminogen activator (t-PA), even though it exhibits only 23% sequence identity with t-PA. This study shows that TSV-PA, t-PA, and batroxobin are quite different in their specificity toward small chromogenic substrates, TSV-PA being less selective than t-PA, and batroxobin not being efficient at all. The specificity of TSV-PA, with respect to t-PA and batroxobin, was investigated further by site-directed mutagenesis in the 189-195 segment, which forms the basement of the S(1) pocket of TSV-PA and presents a His at position 192 and a unique Phe at position 193. This study demonstrates that Phe(193) plays a more significant role than His(192) in determining substrate specificity and inhibition resistance. Interestingly, the TSV-PA variant F193G possesses a 8-9-fold increased activity for plasminogen and becomes sensitive to bovine pancreatic trypsin inhibitor.

Self-association and domains of interactions of an amphipathic helix peptide inhibitor of HIV-1 integrase assessed by analytical ultracentrifugation and NMR experiments in trifluoroethanol/H(2)O mixtures.

EAA26 (VESMNEELKKIIAQVRAQAEHLKTAY) is a better inhibitor of human immunodeficiency virus, type 1, integrase than its parent Lys-159, reproducing the enzyme segment 147-175 with a nonpolar-polar/charged residue periodicity defined by four helical heptads (abcdefg) prone to collapse into a coiled-coil. Circular dichroism, nuclear magnetic resonance, sedimentation equilibrium, and chemical cross-linking were used to analyze EAA26 in various trifluoroethanol/H(2)O mixtures. In pure water the helix content is weak but increases regularly up to 50-60% trifluoroethanol. In contrast the multimerization follows a bell-shaped curve with monomers in pure water, tetramers at 10% trifluoroethanol, and dimers at 40% trifluoroethanol. All suggest that interhelical interactions between apolar side chains are required for the coiled-coil formation of EAA26 and subsist at medium trifluoroethanol concentration. The N(H) temperature coefficients measured by nuclear magnetic resonance show that at low trifluoroethanol concentration the amide groups buried in the hydrophobic interior of four alpha-helix bundles are weakly accessible to trifluoroethanol and are only weakly subject to its hydrogen bond strengthening effect. The increased accessibility of trifluoroethanol to buried amide groups at higher trifluoroethanol concentration entails the reduction of the hydrophobic interactions and the conversion of helix tetramers into helix dimers, the latter displaying a smaller hydrophobic interface. The better inhibitory activity of EAA26 compared with Lys-159 could arise from its better propensity to form a helix bundle structure with the biologically important helical part of the 147-175 segment in integrase.

Molecular modeling of an active loop structure in lysozyme. Sequence effects or crystal packing?

The Val99-Gly 104 variable region in egg white lysozyme is part of the active site cleft and of the epitope recognized by some monoclonal antibodies. In general, this loop is found in a conformation inflected towards the active site (proximal conformational) such as in free hen lysozyme (HEL). But in a lysozyme such as Japanese quail's (JEL), the loop turns away from the active site cleft (distal conformation). In order to differentiate sequence effects from crystal packing, we generated and refined loop conformations for the 99-104 variable region in lysozyme, then estimated their relative conformational free energies. Some of the results indicate that (i) the flexibility of the 99-104 segment is much greater for HEL than for JEL sequences when unconstrained by the crystal lattice, (ii) for JEL, only distal structures are favored, while for HEL the states span the zone between proximal and distal regions, and (iii) epitopes elucidated from crystal structures may not always be conserved in solution. For the JEL loop, model building shows that an energy-costly distal to proximal transition appears necessary. Finally, analysis of available structural data indicates that changes of humidity, temperature and pressure on loop conformation are negligible.

Conformational aspects of HIV-1 integrase inhibition by a peptide derived from the enzyme central domain and by antibodies raised against this peptide.

Monospecific antibodies were raised against a synthetic peptide K159 (SQGVVESMNKELKKIIGQVRDQAEHLKTA) reproducing the segment 147-175 of HIV-1 integrase (IN). Synthesis of substituted and truncated analogs of K159 led us to identify the functional epitope reacting with antibodies within the C-terminal portion 163-175 of K159. Conformational studies combining secondary structure predictions, CD and NMR spectroscopy together with ELISA assays, showed that the greater is the propensity of the epitope for helix formation the higher is the recognition by anti-K159. Both the antibodies and the antigenic peptide K159 exhibited inhibitory activities against IN. In contrast, neither P159, a Pro-containing analog of K159 that presents a kink around proline but with intact epitope conformation, nor the truncated analogs encompassing the epitope, were inhibitors of IN. While the activity of antibodies is restricted to recognition of the sole epitope portion, that of the antigenic K159 likely requires interactions of the peptide with the whole 147-175 segment in the protein [Sourgen F., Maroun, R.G., Frère, V., Bouziane, A., Auclair, C., Troalen, F. & Fermandjian, S. (1996) Eur. J. Biochem. 240, 765-773]. Actually, of all tested peptides only K159 was found to fulfill condition of minimal number of helical heptads to achieve the formation of a stable coiled-coil structure with the IN 147-175 segment. The binding of antibodies and of the antigenic peptide to this segment of IN hampers the binding of IN to its DNA substrates in filter-binding assays. This appears to be the main effect leading to inhibition of integration. Quantitative analysis of filter-binding assay curves indicates that two antibody molecules react with IN implying that the enzyme is dimeric within these experimental conditions. Together, present data provide an insight into the structure-function relationship for the 147-175 peptide domain of the enzyme. They also strongly suggest that the functional enzyme is dimeric. Results could help to assess models for binding of peptide fragments to IN and to develop stronger inhibitors. Moreover, K159 antibodies when expressed in vivo might exhibit useful inhibitory properties.

Structural basis of the gp120 superantigen-binding site on human immunoglobulins.

B cell superantigens (SAg) interact with normal human nonimmune Igs (Igs), independently of the light chain isotype, and activate a large proportion of the B cell repertoire. Recently, the major envelope protein of HIV-1, gp120, was found to exhibit SAg-like properties for B cells with potential pathologic consequences for the infected host. This unconventional mode of interaction contrasts with its binding to immunization-induced Abs, which requires the tertiary structure of the heavy and light chain variable regions. In this report, we have examined the structural basis of the interaction between human Igs and gp120. We found that gp120 binding is restricted to Igs from the V(H)3 gene family and that the two V(H) genes 3-23 and 3-30, known to be overutilized during all stages of B cell development, frequently impart gp120 binding. We also provide evidence that the viral gp120 SAg can interact with only a subset of the human V(H)3+ Igs that can convey binding to the prototypic bacterial B cell SAg protein A from Staphylococcus aureus. Finally, we have identified amino acid positions present primarily in the first and third framework regions of the Ig heavy chain variable region, outside the conventional hypervariable loops, which correlate with gp120 binding. In a three-dimensional sequence-homology model, these residues partially overlap with the predicted SAg protein A binding site for V(H)3+ Igs.

Comparative structural analysis by [1H,31P]-NMR and restrained molecular dynamics of two DNA hairpins from a strong DNA topoisomerase II cleavage site.

The structural analysis of two single-stranded DNAs d(AGCTTATCATCGATAAGCT) (ATC-19) and d(AGCTTATCGATGATAAGCT) (GAT-19) was performed by NMR and restrained molecular dynamics. These oligonucleotides reproduce the 15-33 segment of phage pBR322 DNA, which contains a strong cleavage site for topoisomerase II coupled to the antitumor drugs VP-16 and ellipticine. Because of their partial palindromic nature, the two oligonucleotides ATC-19 and GAT-19 may fold back into stable hairpin structures, consisting of a stem of eight base-pairs and a loop of three residues. NMR assignments and conformational parameters were determined from combined 2D NOESY, COSY and 1H-31P spectra. Conformations of ATC-19 and GAT-19 hairpins were calculated using the X-PLOR 3.1 program. Structures were generated through simulated annealing procedures starting from 50 structures with randomized torsion angles. A good convergence was observed for ATC-19 molecules, while no consensus was found for GAT-19. Within the GAT-19 loop, the base stacking was poor and no hydrogen bond could be detected. In contrast, ATC-19 displayed a well-defined three residue loop stabilized by both extensive base stackings and hydrogen bonding between the N3 atom of the adenine ring and the amino group of the cytosine ring. The results confirm our earlier ATC-19 structure obtained by a completely different calculation procedure (JUMNA) and the higher thermal stability of ATC-19 compared to GAT-19. Moreover, due to its mismatched base-pair, the ATC-19 loop may be better described as a single residue loop rather than a three residue loop. Comparison of this loop to those containing sheared purine.purine base-pairs revealed striking resemblances, particularly on the backbone angle combination. Finally, the differences observed between the ATC-19 and GAT-19 structures could help toward understanding the sequential cleavage of DNA strands by topoisomerase II.

Helical and coiled-coil-forming properties of peptides derived from and inhibiting human immunodeficiency virus type 1 integrase assessed by 1H-NMR--use of NH temperature coefficients to probe coiled-coil structures.

Human immunodeficiency virus type 1 integrase (HIV-1 IN) which catalyzes viral DNA integration into the host genome of infected cells represents an attractive target for AIDS therapy. We have previously demonstrated the ability of the IN-(147-175)-peptide derived from the catalytic core domain of HIV-1 IN to inhibit the enzyme activity in vitro. IN-(147-175)-peptide contains four heptad repeats and displays a high propensity for coiled-coil formation while its [P159]IN-(147-175)-peptide analog (Lys159-->Pro in the protein, Lys13-->Pro in the peptide) is unable to form a stable coiled-coil and is devoid of inhibitory activity [Sourgen, F., Maroun, R. G., Frère, V., Bouziane, M., Auclair, C., Troalen, F. & Fermandjian, S. (1996) Eur. J. Biochem. 240, 765-773]. Now, we report results from an NMR study on IN-(147-175)-peptide and [P159]IN-(147- 175)-peptide as well as on an optimized [E156, A163, A167]IN-(147-175)-peptide that is a better inhibitor of IN than IN-(147-175)-peptide. While in aqueous solution, IN-(147-175)-peptide and [P159]IN-(147-175)-peptide display only nascent helical features, [E156, A163, A167]IN-(147-175)-peptide exhibits 20% of helical content. In 20% trifluoroethanol/80% H2O, the helix content is the highest for [E156, A163, A167]IN-(147-175)-peptide (approximately 70%) and the lowest for [P159]IN-(147-175)-peptide (approximately 40%), due to a local helix break caused by the Pro residue. The NHs of residues in the two central helical heptads (a-g) of IN-(147-175)-peptide and [E156, A163, A167]IN-(147-175)-peptide display a regular periodic variation of their temperature coefficients in 20% trifluoroethanol. The b, c and f residues on the hydrophilic face of the amphipathic helix show high coefficients reflecting hydrogen bonded NHs, while the a and d residues on the hydrophobic face exhibit low coefficients, near random-coil values. The particular arrangement of the hydrophobic side-chains of a and d residues at the coiled-coil interface reduces the access of trifluoroethanol molecules to their amide groups. The inability of trifluoroethanol molecules to create interactions with the amide C=O groups, these being required to strengthen the intrahelical C=O...H-N hydrogen bonds, is the main cause for observation of heptadic a and d residues with low NH temperature coefficients. Such effects concern mostly the two central helical heptads of IN-(147-175)-peptide and [E156, A163, A167]IN-(147-175)-peptide implying that these ones are engaged in stable parallel coiled coils. Our results provide a link between the propensity of peptides for helix formation, their coiled-coil properties and their efficiency to inhibit IN.