Fr Francium

Cellular redox conditions affect Gsp amidase activity in Escherichia coli. Guided by the structure and catalytic mechanism of the amidase, we designed and synthesized an acyloxymethyl ketone-based activity probe containing a biotin handle. This probe was used to monitor Gsp amidase activity in E. coli lysates that had been subjected to oxidative or methylglyoxal-induced stress.

Sponge natural product biosynthesis: A highly sensitive in vivo protocol based on ¹⁴C radiolabeled precursors and beta-imager autoradiography allowed the unraveling of the origin of the pyrrole 2-aminoimidazole-containing key biosynthetic intermediate oroidin. Proline and lysine are now proposed as the early precursors of the pyrrole and the 2-aminoimidazole moieties of oroidin respectively.

The majority of studies on DNA triple helices have been focused on pH-sensitive parallel triplexes with Hoogsteen CT-containing third strands that require protonation of cytosines. Reverse Hoogsteen GT/GA-containing antiparallel triplex-forming oligonucleotides (TFOs) do not require an acidic pH but their applicability in triplex technology is limited because of their tendency to form undesired highly stable aggregates such as G-quadruplexes. In this study, G-rich oligonucleotides containing 2–4 insertions of twisted intercalating nucleic acid (TINA) monomers are demonstrated to disrupt the formation of G-quadruplexes and form stable antiparallel triplexes with target DNA duplexes. The structure of TINA-incorporated oligonucleotides was optimized, the rules of their design were established and the optimal triplex-forming oligonucleotides were selected. These oligonucleotides show high affinity towards a 16 bp homopurine model sequence from the HIV-1 genome; dissociation constants as low as 160 nM are observed whereas the unmodified TFO does not show any triplex formation and instead forms an intermolecular G-quadruplex with T_m exceeding 90 °C in the presence of 50 mM NaCl. Here we present a set of rules that help to reach the full potential of TINA-TFOs and demonstrate the effect of TINA on the formation and stability of triple helical DNA.

Want to form an antiparallel DNA triplex? It's easy! A TG-DNA sequence with a run of six guanines exists as a stable tetramolecular parallel G-quadruplex and does not bind to the target polypurine tract of HIV-1 proviral DNA. TINA modification converts this sequence into an efficient and sequence-selective triplex-forming oligonucleotide with the dissociation constant as low as 160 nM.

Light-triggered motor: Incorporation of a photocleavable group onto a phosphoserine residue of the regulator of a motor protein allows light-induced activation with spatial and temporal precision inside a living cell.

Now found in bacteria: An increasing number of genome sequences indicate that bacteria possess a variety of terpenoid cyclase genes. The characterization of two sesquiterpene cyclase (SC) genes found in the draft genome sequence of Streptomyces citricolor is described here. Our study strongly supports the idea that genome mining is a useful approach in revealing the terpenoid diversity in bacteria.

Order in disorder: The characterization of intrinsically disordered proteins by NMR spectroscopy is a necessity on the one hand and a continuous challenge on the other. We propose two experiments that provide diagnostic parameters to monitor the degree of unfolding of a polypeptide. The test was performed on the yeast Cox17 protein, known to gain its function through maturation from an intrinsically disordered state (see figure).

Modes of transport: A leucine-zipper-tagged GFP was transported into cells by “zipping” it (red) to it's complementary leucine zipper (blue) functionalized with a cell-penetrating peptide (CPP). This transport system has an inherent modularity as the CPP is “clicked” to the leucine zipper, and then noncovalently bound to the protein, thus making it system particularly useful for targeting studies.

Production of typical plant metabolites by a fungal enzyme: Fungal prenyltransferases of the DMATS superfamily are mainly involved in the biosynthesis of prenylated indole alkaloids, but also catalyze the prenylation of tyrosine and naphthalene derivatives. In this study, nine prenylated flavonoids were produced by using the recombinant dimethylallyltryptophan synthase 7-DMATS from Aspergillus fumigatus.

Chain armor against tumor cells: The oxazole side chain in the antimitotic agent rhizoxin S2 (1) was successfully replaced through mutasynthesis by using an engineered mutant impaired in heterocyclization. Incorporation of 12 non-natural surrogates into fully processed rhizoxin analogues revealed a remarkable substrate flexibility of the PKS–NRPS hybrid.

Tailoring guide: We have used structure-guided saturation mutagenesis followed by colorimetric screening to identify mutant malonyl-CoA synthetases with altered substrate specificity. One particular mutant displayed a 240-fold shift in specificity (see graphic). These mutant enzymes will be useful tools for providing extender units to probe the activity of polyketide synthases.