Project description:Phosphinothricin tripeptide (PTT, phosphinothricylalanylalanine) is a natural-product antibiotic and potent herbicide that is produced by Streptomyces hygroscopicus ATCC 21705 (ref. 1) and Streptomyces viridochromogenes DSM 40736 (ref. 2). PTT has attracted widespread interest because of its commercial applications and unique phosphinic acid functional group. Despite intensive study since its discovery in 1972 (see ref. 3 for a comprehensive review), a number of steps early in the PTT biosynthetic pathway remain uncharacterized. Here we report a series of interdisciplinary experiments involving the construction of defined S. viridochromogenes mutants, chemical characterization of accumulated intermediates, and in vitro assay of selected enzymes to examine these critical steps in PTT biosynthesis. Our results indicate that early PTT biosynthesis involves a series of catalytic steps that to our knowledge has not been described so far, including a highly unusual reaction for carbon bond cleavage. In sum, we define a pathway for early PTT biosynthesis that is more complex than previously appreciated.
Project description:Polymeric nanocarriers have been extensively used in medicinal applications for drug delivery. However, intravenous nanocarriers circulating in the blood will be rapidly cleared from the mononuclear macrophage system. The surface physicochemical characterizations of nanocarriers are the primary factors to determine their fate in vivo, such as evading the reticuloendothelial system, exhibiting long blood circulation times, and accumulating in the targeted site. In this work, we develop a series of polyurethane micelles containing segments of an anionic tripeptide, hydrophilic mPEG, and disulfide bonds. It is found that the long hydrophilic mPEG can shield the micellar surface and have a synergistic effect with the negatively charged tripeptide to minimize macrophage phagocytosis. Meanwhile, the disulfide bond can rapidly respond to the intracellular reduction environment, leading to the acceleration of drug release and improvement of the therapeutic effect. Our results verify that these anionic polyurethane micelles hold great potential in the development of the stealth immune system and controllable intracellular drug transporters.
Project description:Secreted amyloid precursor protein-alpha (sAPPα), generated by enzymatic processing of the APP, possesses a range of neurotrophic and neuroprotective properties and plays a critical role in the molecular mechanisms of memory and learning. One of the key active regions of sAPPα is the central APP domain (E2) that contains within it the tripeptide sequence, RER. This sequence is exposed on the surface of a coiled coil substructure of E2. RER has by itself displayed memory-enhancing properties, and can protect newly formed engrams from interference in a manner similar to that displayed by sAPPα itself. In order to determine whether RER mimics other properties of sAPPα, we investigated the electrophysiological effects of the N-terminal protected acetylated RER (Ac-RER) and an isoform containing a chiral switch in the first amino acid from an l- to a d-orientation (Ac-rER), on synaptic plasticity. We found that, like sAPPα, exogenous perfusion with nanomolar concentrations of Ac-RER or Ac-rER enhanced the induction and stability of long-term potentiation (LTP) in area CA1 of rat and mouse hippocampal slices, in a protein synthesis- and trafficking-dependent manner. This effect did not occur with a control Ac-AAA or Ac-IFR tripeptide, nor with a full-length sAPPα protein where RER was substituted with AAA. Ac-rER also protected LTP against amyloid-beta (Aβ25 - 35)-induced LTP impairment. Our findings provide further evidence that the RER-containing region of sAPPα is functionally significant and by itself can produce effects similar to those displayed by full length sAPPα, suggesting that this tripeptide, like sAPPα, may have therapeutic potential.
Project description:The late stages of biosynthesis of phosphinothricin tripeptide (PTT) involve peptide formation and methylation on phosphorus. The exact timing of these transformations is not known. To provide insight into this question, we developed a heterologous expression system for PhsA, one of three NRPS proteins in PTT biosynthesis. The apparent k(cat)/K(m) value for ATP-pyrophosphate exchange activity for d,l-N-acetylphosphinothricin was 3.5 muM(-1) min(-1), whereas the k(cat)/K(m,app) for l-N-acetyldemethylphosphinothricin was 0.5 microM(-1) min(-1), suggesting the former might be the physiological substrate. Each substrate could be loaded onto the phosphopantetheine arm of the thiolation domain as observed by Fourier transform mass spectrometry (FTMS).
Project description:Unlike linear peptides, analysis of cyclic peptides containing disulfide bonds is not straightforward and demands indirect methods to achieve a rigorous proof of structure. Three peptides that belong to this category, p-Cl-Phe-DPDPE, DPDPE, and CTOP, were analyzed and the results are presented in this paper. The great potential of two dimensional NMR and ESI tandem mass spectrometry was harnessed during the course of peptide characterizations. A new RP-HPLC method for the analysis of trifluoroacetic acid is also presented. It is robust, simple, and efficient compared to the currently available methods.
Project description:An enantioselective domino Michael-Michael reaction of nitroolefins and 2-nitro-3-arylacrylates has been established, which provided a series of spirocyclopentane oxindoles with four consecutive stereocenters including quaternary α-nitro esters with good yields (up to 73%) and excellent enantioselectivities (up to 97% ee). The reaction was realized and optimized with the aid of a chiral squaramide-amine catalyst. The structures of 11 products were confirmed by single-crystal X-ray diffraction analysis.
Project description:Identifying oncological applications for drugs that are already approved for other medical indications is considered a possible solution for the increasing costs of cancer treatment. Under the hypothesis that nutritional stress through fasting might enhance the antitumour properties of at least some non-oncological agents, by screening drug libraries, we find that cholesterol biosynthesis inhibitors (CBIs), including simvastatin, have increased activity against cancers of different histology under fasting conditions. We show fasting's ability to increase CBIs' antitumour effects to depend on the reduction in circulating insulin, insulin-like growth factor-1 and leptin, which blunts the expression of enzymes from the cholesterol biosynthesis pathway and enhances cholesterol efflux from cancer cells. Ultimately, low cholesterol levels through combined fasting and CBIs reduce AKT and STAT3 activity, oxidative phosphorylation and energy stores in the tumour. Our results support further studies of CBIs in combination with fasting-based dietary regimens in cancer treatment and highlight the value of fasting for drug repurposing in oncology.
Project description:In recent years it has become apparent that aminoacyl-tRNAs are not only crucial components involved in protein biosynthesis, but are also used as substrates and amino acid donors in a variety of other important cellular processes, ranging from bacterial cell wall biosynthesis and lipid modification to protein turnover and secondary metabolite assembly. In this review, we focus on tRNA-dependent biosynthetic pathways that generate modified cyclic dipeptides (CDPs). The essential peptide bond-forming catalysts responsible for the initial generation of a CDP-scaffold are referred to as cyclodipeptide synthases (CDPSs) and use loaded tRNAs as their substrates. After initially discussing the phylogenetic distribution and organization of CDPS gene clusters, we will focus on structural and catalytic properties of CDPSs before turning to two recently characterized CDPS-dependent pathways that assemble modified CDPs. Finally, possible applications of CDPSs in the rational design of structural diversity using combinatorial biosynthesis will be discussed before concluding with a short outlook.
Project description:We report the design, synthesis, membrane activity, biophysical characterization, and in vitro antibacterial activities of cationic cyclic D,L-alpha-glycopeptides bearing d-glucosamine (GlcNH(2)), D-galactose (Gal), or D-mannose (Man) glycosyl side chains.
Project description:In the title compound, C(12)H(15)NO(7)·0.5H(2)O, there are two independent mol-ecules in the asymmetric unit, together with one water molecule. The pyran-oside rings each have close to a (1)C(4) chair conformation and the nitro groups are almost coplanar with the benzene rings. The water mol-ecule links the two independent mol-ecules through O-H⋯O hydrogen bonds. All the hydroxyl groups are involved in hydrogen-bond inter-actions, giving rise to a three-dimensional network.