SB056 is a novel semi-synthetic antimicrobial peptide with a dimeric dendrimer scaffold. Active against both Gram-negative and -positive bacteria, its mechanism has been attributed to a disruption of bacterial membranes. The branched peptide was shown to assume a β-stranded conformation in a lipidic environment. Here, we report on a rational modification of the original, empirically derived linear peptide sequence [WKKIRVRLSA-NH2, SB056-lin]. We interchanged the first two residues [KWKIRVRLSA-NH2, β-SB056-lin] to enhance the amphipathic profile, in the hope that a more regular β-strand would lead to a better antimicrobial performance. MIC values confirmed that an enhanced amphiphilic profile indeed significantly increases activity against both Gram-positive and -negative strains. The membrane binding affinity of both peptides, measured by tryptophan fluorescence, increased with an increasing ratio of negatively charged/zwitterionic lipids. Remarkably, β-SB056-lin showed considerable binding even to purely zwitterionic membranes, unlike the original sequence, indicating that besides electrostatic attraction also the amphipathicity of the peptide structure plays a fundamental role in binding, by stabilizing the bound state. Synchrotron radiation circular dichroism and solid-state 19F-NMR were used to characterize and compare the conformation and mobility of the membrane bound peptides. Both SB056-lin and β-SB056-lin adopt a β-stranded conformation upon binding POPC vesicles, but the former maintains an intrinsic structural disorder that also affects its aggregation tendency. Upon introducing some anionic POPG into the POPC matrix, the sequence-optimized β-SB056-lin forms well-ordered β-strands once electro-neutrality is approached, and it aggregates into more extended β-sheets as the concentration of anionic lipids in the bilayer is raised. The enhanced antimicrobial activity of the analogue correlates with the formation of these extended β-sheets, which also leads to a dramatic alteration of membrane integrity as shown by 31P-NMR. These findings are generally relevant for the design and optimization of other membrane-active antimicrobial peptides that can fold into amphipathic β-strands.
Caves are a particular extreme environment for humans because of the high humidity, muddy, darkness, and slippery conditions. Moreover, explorations can last many hours or even days, and require extensive climbing and ropework. Thus, given the uniqueness of the environment and the duration of effort, caving is a peculiar sport activity. However, very little is known about physical capacity of cavers as well as about their energy expenditure (EE) during caving. The physical capacity of 17 (7 females) expert cavers (age 43.9±7.3 yrs, body mass 69.8±15.4 kg, height 167.9±10.9 cm) was assessed during an incremental cycle-ergometer test with gas exchange analysis (CPX). Moreover, a wearable metabolic band (Armband Fit Core), equipped with sensors measuring skin temperature, heat flux, galvanic skin response and acceleration, was used to estimate their EE during caving. In terms of physical capacity, the CPX test showed that cavers had a maximum oxygen uptake (VO2max) of 2248.7±657.8 ml*min−1 (i.e. 32.4±6.4 ml/kg*min−1), while anaerobic threshold (AT) occurred on average at 74.5 % of VO2max. Results from caving sessions provided an average time spent in cave of 9.4±1.2 hours while the average EE was 268.8±54.8 Kcal/h, which corresponded to about 40% of the maximum aerobic capacity measured by the CPX. A mean distance of 10.6±2.2 km was covered by subjects. Data from the present investigation provide evidence that cavers have a low level of aerobic physical capacity, which is only slightly higher than that of sedentary people, thereby suggesting that a high aerobic fitness is not needed by cavers. Moreover, during caving the EE was on average well below the level of AT. However, in absolute terms, the total EE was elevated (i.e. 2672.3±576 Kcal in total) due to the long time spent in caving. These facts should be taken into account when preparing training as well as diet programs for these athletes. Support or Funding Information This study was supported by the University of Cagliari and the Italian Ministry of Scientific Research. mean values (±SD) of oxygen uptake (VO2) at maximum workload (VO2max), at anaerobic threshold (VO2AT), and during caving (VO2caving). † = p<0.05 vs. VO2AT, * = p<0.05 vs. VO2caving
Abstract Halimium is a genus of Cistaceae , containing a small group of shrub species found in open vegetation types and in degraded forest patches throughout the western and central Mediterranean region. We recently described the morpho-anatomical features of the ectomycorrhizae formed by Scleroderma meridionale on Halimium halimifolium , but the mycorrhizal biology of this host plant genus is still largely unexplored. Here, we report new data on the ectomycorrhizal fungal symbionts of Halimium , based on the collection of sporocarps and ectomycorrhizal root tips in pure stands occurring in Sardinia, Italy. To obtain a broader view of Halimium mycorrhizal and ecological potential, we compiled a comprehensive and up-to-date checklist of fungal species reported to establish ectomycorrhizae on Halimium spp. on the basis of field observations, molecular approaches, and mycorrhiza synthesis. Our list comprises 154 records, corresponding to 102 fungal species and 35 genera, revealing a significant diversity of the Halimium ectomycorrhizal mycobiota. Key ectomycorrhizal genera like Russula , Lactarius / Lactifluus , Amanita , Inocybe , and Cortinarius account for more than half of all mycobionts. A large proportion of Halimium fungal species are shared with other host plants in various ecological settings, suggesting a critical role of common mycorrhizal networks in the function played by this shrub in various Mediterranean ecosystems.
Purified lentil seedling amino oxidase (LSAO) is homogeneous in the analytical ultracentrifuge, but shows heterogeneity in gel-filtration HPLC and in PAGE. Two components were obtained from HPLC and PAGE, but at least 6-7 subforms were seen by electrofocusing techniques. The chromatographic and electrophoretic forms are not interconvertible, indicating the presence of covalent differences. The electrophoretic pattern, but not the chromatographic pattern, is modified by treating the enzyme with a pool of glycohydrolases. The copper-free enzyme shows the same type of heterogeneity as the native enzyme, this ruling out the possibility that some subforms were due to the presence of apoenzyme.
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