Alberto Sutera

Photosynthesized silver–polyaminocyclodextrin nanocomposites as promising antibacterial agents with improved activity

Ag nanocomposites were prepared by photoreduction of ammoniacal silver acetate in the presence of poly-{6-[3-(2-(3-aminopropylamino)ethylamino)propylamino]}-(6-deoxy)-β-CD (amCD). The obtained systems were characterized by means of various complementary techniques (UV-vis, FT-IR, TEM, SAED). In particular, FT-IR spectroscopy evidenced a partial oxidative degradation of the polyamine branches of the capping auxiliary, due to the fact that these groups function as a sacrificial reducing agent in the photoinduced formation of the Ag metal core. TEM and SAED micrographs showed that the Ag cores possess a relatively low polydispersity and a significantly crystalline character. The Ag–amCD systems were assayed for antibacterial activity, using Escherichia coli and Kocuria rhizophila as Gram-negative and Gram-positive tester strains respectively. In addition, the systems function as supramolecular drug carriers, able to bind the β-lactam antibiotic ampicillin, as demonstrated by polarimetric measurements. Antimicrobial assays revealed MIC90 values against E. coli and K. rhizophila as large as a 5 and 1 μg mL−1 respectively. Moreover, the interaction of the Ag–amCD with ampicillin resulted in a synergistic improvement of the antibacterial activity. This study provides insights on the attractive possibility to use a photochemical methodology to produce bioactive supramolecular systems to be employed as powerful and tunable antimicrobial agents.

The effects of structural changes on the anti-microbial and anti-proliferative activities of diimidazolium salts

An array of diimidazolium salts has been synthesized and used to investigate their anti-microbial and anti-proliferative activities. In particular, salts based on the 3,3′-di-n-alkyl-1,1′-(1,n-phenylenedimethylene)diimidazolium cation and differing in the alkyl chain length on the imidazolium ion, the isomeric substitution on the aromatic spacer and in the anion nature were used. The anti-proliferative activity was evaluated against cervical (HeLa), colon adenocarcinoma (HT-29) and breast (SKBR3) cancer cell lines. In the latter case, also a morphological assessment after treatment with salts was performed. All salts were tested for their hemolytic activity against human erythrocytes. On the other hand, the anti-microbial activity was investigated using both Gram-negative (Escherichia coli) and Gram-positive (Kokuria rhizophila, Staphilococcus aureus and Bacillus subtilis) strains. On the whole, the data collected demonstrate that biological activity is the result of the combined action of both cation and anion structures. In general, the cation hydrophobicity plays the most significant role with structural features of the anion becoming more relevant in the presence of a shorter alkyl chain on the cationic head.

TrpM, a Small Protein Modulating Tryptophan Biosynthesis and Morpho-Physiological Differentiation in Streptomyces coelicolor A3(2).

In the model actinomycete Streptomyces coelicolor A3(2), small open reading frames encoding proteins with unknown functions were identified in several amino acid biosynthetic gene operons, such as SCO2038 (trpX) in the tryptophan trpCXBA locus. In this study, the role of the corresponding protein in tryptophan biosynthesis was investigated by combining phenotypic and molecular analyses. The 2038KO mutant strain was characterized by delayed growth, smaller aerial hyphae and reduced production of spores and actinorhodin antibiotic, with respect to the WT strain. The capability of this mutant to grow on minimal medium was rescued by tryptophan and tryptophan precursor (serine and/or indole) supplementation on minimal medium and by gene complementation, revealing the essential role of this protein, here named TrpM, as modulator of tryptophan biosynthesis. His-tag pull-down and bacterial adenylate cyclase-based two hybrid assays revealed TrpM interaction with a putative leucyl-aminopeptidase (PepA), highly conserved component among various Streptomyces spp. In silico analyses showed that PepA is involved in the metabolism of serine, glycine and cysteine through a network including GlyA, CysK and CysM enzymes. Proteomic experiments suggested a TrpM-dependent regulation of metabolic pathways and cellular processes that includes enzymes such as GlyA, which is required for the biosynthesis of tryptophan precursors and key proteins participating in the morpho-physiological differentiation program. Altogether, these findings reveal that TrpM controls tryptophan biosynthesis at the level of direct precursor availability and, therefore, it is able to exert a crucial effect on the morpho-physiological differentiation program in S. coelicolor A3(2).

Supramolecular Hydro- and Ionogels: A Study of Their Properties and Antibacterial Activity

Diimidazolium-based organic salts, bearing peptides or amino acids as anions have been synthesised and tested for their gelling ability in biocompatible solvents. These low molecular weight salts were successfully used as gelators in phosphate buffered saline (PBS) solution and ionic liquids. Then, the properties of the obtained soft materials were analysed in terms of melting temperature and gel strength as accounted for by rheological investigations. The gel-phase formation was studied by using UV/Vis and resonance light scattering measurements, whereas the morphology of the soft materials was analysed by using polarised optical microscopy and scanning electron microscopy. To get information about the organisation of the gelator in the gelatinous matrix, X-ray diffraction measurements were performed both on the neat gelators and their gels. The results collected show that the properties of the gel phases, like the thermal stability, the self-repairing ability, the resistance to flow as well as the morphology, are dependent on the nature of the anion. Furthermore, bioassays revealed that the obtained diimidazolium organic salts possessed antimicrobial activity, against gram-negative and gram-positive tester strains. In particular and noteworthy, the diimidazolium organic salts exert a bactericidal capability, which was retained even if they are included in the gel phase. Thus, a novel kind of bioactive soft material was obtained that could be fruitfully employed as a non-covalent coating exerting antibacterial capability.

Pirin: A novel redox-sensitive modulator of primary and secondary metabolism in Streptomyces

Pirins are evolutionarily conserved iron-containing proteins that are found in all kingdoms of life, and have been implicated in diverse molecular processes, mostly associated with cellular stress. In the present study, we started from the evidence that the insertional inactivation of pirin-like gene SAM23877_RS18305 (pirA) by ΦC31 Att/Int system-based vectors in spiramycin-producing strain Streptomyces ambofaciens ATCC 23877 resulted in marked effects on central carbon and energy metabolism gene expression, high sensitivity to oxidative injury and repression of polyketide antibiotic production. By using integrated transcriptomic, proteomic and metabolite profiling, together with genetic complementation, we here show that most of these effects could be traced to the inability of the pirA-defective strain to modulate beta-oxidation pathway, leading to an unbalanced supply of precursor monomers for polyketide biosynthesis. Indeed, in silico protein-protein interaction modeling and in vitro experimental validation allowed us to demonstrate that PirA is a novel redox-sensitive negative modulator of very long-chain acyl-CoA dehydrogenase, which catalyzes the first committed step of the beta-oxidation pathway.