1,721,166 research outputs found
Protein folding and aggregation: metals and biomolecules in protein conformational diseases
No full textOptimizing a kinase assay for IKK.beta. on an HTS station
No full textUsing a commercially available time-resolved fluorescence resonance energy transfer (TR-FRET)—based assay for IKKβ, the authors have automated the assay procedure on a high-throughput screening station to carry out screening campaigns on multiwell plates. They have determined the Z' factor and optimized volumes, times, and time-resolved fluorescence parameters. They have also compared 2 kinases with different fusion tags, the influence of different enzyme/substrate ratios and of DMSO presence at different concentration. The authors found that glutathione S-transferase (GST)—fused IKKβ shows better signal-to-noise (S/N) ratios over the poly-histidine-tagged variant. The substrate can be used at 50 nM with optimal performances when the enzyme is used at 2 nM. DMSO at 0.2% and 1% only slightly affects the S/N ratio, whereas when used at 2%, the final concentration deriving from a 50-fold dilution from a 5-mM stock solution in pure solvent, S/N undergoes a decrease of about 15%. Under the optimized conditions, the assay Z' factor calculated over 192 data points has an optimized value of 0.881 and allows the testing of 94 molecules in quadruplicate in 140 min
Phytocannabinoids and cannabimimetic drugs: Recent patents in central nervous system disorders
No full textAgents acting via cannabinoid receptors have been widely developed; starting from the chemical structure of phytocannabinoids isolated from cannabis sativa plant, specific and selective compounds of these receptors have been produced ranging from partial to full agonists and /or antagonists endowed with different potency. The enhanced interest on developing such classes of drugs is due to the beneficial properties widely reported by both anecdotal reports and scientific studies describing the potential medicinal use of cannabinoids and their derivatives in numerous pathological conditions in both in vitro and in vivo models. The use of these drugs has been found to be of benefit in a wide number of neurological and neuropsychiatric disorders, and in many other diseases ranging from cancer, atherosclerosis, stroke, hypertension, inflammatory related disorders, and autoimmune diseases, just to mention some. In particular, being the cannabinoid CB1 receptor a central receptor expressed by neurons of the central nervous system, the attention for the treatment of neurological diseases has been mainly focused on compounds acting via this receptor, however some of these compounds has been showed to act by alternative pathways in some cases unrelated to CB1 receptors. Nonetheless, endocannabinoids are potent regulators of the synaptic function in the central nervous system and their levels are modulated in neurological diseases. In this study, we focused on endocannabinoid mechanism of action in neuronal signaling and on cannabimimetic drug potential application in neurological disorders. Finally, novel patents on cannabis-based drugs with applicability in central nervous system disorders are highlighted, to suggest future potential therapeutic utility of derivatives of this ancient plant
Targeting "Undruggable" Proteins: Design of Synthetic Cyclopeptides
No full textThe development of synthetic macrocycles represents a powerful approach toward the identification of new protein binders or inhibitors of Protein-Protein Interactions (PPI) which are known to play key biological roles in cancer signaling as well as in the regulation of cell division cycle. Structural investigations led to identify "hot loops" sharing common motifs that are mainly involved in PPIs. Most PPIs occur through large and flat surfaces; currently these protein complexes are defined as "undruggable" by conventional drug-discovery approaches, since the identification of small molecules to inhibit these targets is often unreachable. Typically macrocycles are 500-2000 Da in size, having 12-membered, or more, ring architecture: they do not obey the Lipinski's rule but, for them nature offers many examples as therapeutic agents such as erythromycin (antibiotic), cyclosporin (immunosuppressant) and somatostatin (hormone). Peptide-based macrocycles offer the advantages of directly mimicking secondary structures involved in PPIs and their pharmacological application is related to the potential improvement of lead peptides in terms of potency, selectivity, stability and cell permeation. The promising relevance of cyclopeptides prompted to develop new synthetic methods for cyclization: often biotechnological approaches as well as regioselective reactions have been employed to cyclize peptides rapidly and nearly quantitatively. Moreover, different synthetic strategies in peptidomimetics' macrocyclization are actually available based on surrogate peptide bonds or NCL (Native Chemical Ligation) methods. In this review we focus on the most common methods for the preparation of cyclopeptides and interesting applications of the last decade
Oxaliplatin vs. cisplatin: Competition experiments on their binding to lysozyme
No full textThe model protein hen egg white lysozyme was challenged
with Oxaliplatin and Cisplatin. ESI mass spectrometry,
surface plasmon resonance and thermal shift analyses
demonstrate the formation of a bis-platinum adduct, though
in very small amounts. Crystals of the bis-platinum adduct
were obtained using two different preparations and the X-ray
structures solved at 1.85 Ǻ and 1.95 Ǻ resolution. Overall, the
obtained data point out that, under the analyzed conditions,
the two Pt drugs have similar affinities for the protein, but
bind on its surface at two non-overlapping sites. In other
words, these two drugs manifest a significantly different
reactivity with this model protein and do not compete for the
same protein binding sites
The effects of histidine substitution of aromatic residues on the amyloidogenic properties of the fragment 264–277 of nucleophosmin 1
No full text: Histidine (His) plays a key role in mediating protein interactions and its unique side chain determines pH responsive self-assembling processes and thus in the formation of nanostructures. In this study, To identify novel self-assembling bioinspired sequences, we analyzed a series of peptide sequences obtained through the point mutation of aromatic residues of 264-277 fragment of nucleophosmin 1 (NPM1) with single and double histidines. Through several orthogonal biophysical techniques and under different pH and ionic strength conditions we evaluated the effects of these substitutions in the amyloidogenic features of derived peptides. The results clearly indicate that both the type of aromatic mutated residue and its position can have different effect on amyloid-like behaviors. They corroborate the crucial role exerted by Tyr271 in the self-assembling process of CTD of NPM1 in AML mutated form and add novel insights in the accurate investigation of how side chain orientations can determine successful design of innovative bioinspired materials
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