A genomic library of Streptomyces cattleya was screened to isolate a gene cluster encoding enzymes responsible for the production of fluorine-containing metabolites. In addition to the previously described fluorinase FlA which catalyzes the formation of 5'-fluoro-5'-deoxyadenosine from S-adenosylmethionine and fluoride, 11 other putative open reading frames have been identified. Three of the proteins encoded by these genes have been characterized. FlB was determined to be the second enzyme in the pathway, catalyzing the phosphorolytic cleavage of 5'-fluoro-5'-deoxyadenosine to produce 5-fluoro-5-deoxy-D-ribose-1-phosphate. The enzyme FlI was found to be an S-adenosylhomocysteine hydrolase, which may act to relieve S-adenosylhomocysteine inhibition of the fluorinase. Finally, flK encodes a thioesterase which catalyzes the selective breakdown of fluoroacetyl-CoA but not acetyl-CoA, suggesting that it provides the producing strain with a mechanism for resistance to fluoroacetate.

No abstract

Fucoidan, a heparin-like sulfated polysaccharide, is rich in brown algae. It has a wide assortment of protective activities against cancer, for example, induction of hepatocellular carcinoma senescence, induction of human breast and colon carcinoma apoptosis, and impediment of lung cancer cells migration and invasion. However, the anti-metastatic mechanism that fucoidan exploits remains elusive. In this report, we explored the effects of fucoidan on cachectic symptoms, tumor development, lung carcinoma cell spreading and proliferation, as well as expression of metastasis-associated proteins in the Lewis lung carcinoma (LLC) cells-inoculated mice model. We discovered that administration of fucoidan has prophylactic effects on mitigation of cachectic body weight loss and improvement of lung masses in tumor-inoculated mice. These desired effects are attributed to inhibition of LLC spreading and proliferation in lung tissues. Fucoidan also down-regulates expression of matrix metalloproteinases (MMPs), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and vascular endothelial growth factor (VEGF). Moreover, the tumor-bearing mice supplemented with fucoidan indeed benefit from an ensemble of the chemo-phylacticity. The fact is that fucoidan significantly decreases viability, migration, invasion, and MMPs activities of LLC cells. In summary, fucoidan is suitable to act as a chemo-preventative agent for minimizing cachectic symptoms as well as inhibiting lung carcinoma metastasis through down-regulating metastatic factors VEGF and MMPs.

The gene cluster encoding biosynthesis of the clinically important glycopeptide antibiotic teicoplanin has been cloned from Actinoplanes teichomyceticus. Forty-nine putative open reading frames (ORFs) were identified within an 89 kbp genetic locus and assigned roles in teicoplanin biosynthesis, export, resistance, and regulation. Two ORFs, designated orfs 1 and 10*, showed significant homology to known glycosyltransferases. When heterologously expressed in Escherichia coli, these glycosyltransferases were shown to catalyze the transfer of UDP-(N-acetyl)-glucosamine onto, respectively, 3-chloro-beta-hydroxytyrosine-6 (3-Cl-6betaHty) and 4-hydroxyphenylglycine-4 (4Hpg) of the teicoplanin heptapeptide aglycone. The product of another ORF, orf11*, was demonstrated in vitro to transfer n-acetyl-, n-butyryl-, and n-octanoyl-groups from acyl-CoA donors either to a free UDP-aminosugar or to an aminosugar moiety in the teicoplanin pseudoaglycone, thus identifying Orf11* as the key acyltransferase in teicoplanin maturation. These findings should accelerate the combinatorial engineering of new and improved glycopeptide drugs.

No abstract

Japanese encephalitis virus (JEV), a neurotropic flavivirus, is one of the major causes of acute encephalitis in humans. After infection, inflammatory reactions and neurological diseases often develop. Still there are no effective drugs available against virus infection. Recently, extracts of algae have been shown to possess a broad range of biological activities including antivirus activity. In this study, we identified that the sulfated polysaccharide extracts from Ulva lactuca can inhibit JEV infection in Vero cells. Mechanistic studies further revealed that the Ulva sulfated polysaccharide extracts can block virus adsorption and thus make the virus unable to enter cells. The Ulva sulfated polysaccharide extracts also effectively decrease the production of pro-inflammatory cytokines in the JEV-infected primary mixed glia cells. In an animal study, the JEV-infected C3H/HeN mice appeared to have neurobehavioral abnormalities on the fifth day and died on the seventh day post infection. However, the JEV-infected mice pretreated with the Ulva sulfated polysaccharide extracts can delay the onset of hind limb paralysis and thereby prevent mice from death.

In the search for new efficacious antibiotics, biosynthetic engineering offers attractive opportunities to introduce minor alterations to antibiotic structures that may overcome resistance. Dbv29, a flavin-containing oxidase, catalyzes the four-electron oxidation of a vancomycin-like glycopeptide to yield A40926. Structural and biochemical examination of Dbv29 now provides insights into residues that govern flavinylation and activity, protein conformation and reaction mechanism. In particular, the serendipitous discovery of a reaction intermediate in the crystal structure led us to identify an unexpected opportunity to intercept the normal enzyme mechanism at two different points to create new teicoplanin analogs. Using this method, we synthesized families of antibiotic analogs with amidated and aminated lipid chains, some of which showed marked potency and efficacy against multidrug resistant pathogens. This method offers a new strategy for the development of chemical diversity to combat antibacterial resistance.

Phospho- N -acetylmuramoyl-pentapeptide translocase (MraY AA ) from Aquifex aeolicus is the binding target for the nucleotide antibiotic muraymycin D2 (MD2). MraY AA in the presence of the MD2 ligand has been crystallized and released, while the interactions between the ligand and active-site residues remain less quantitatively and qualitatively defined. We characterized theoretically the key residues involved in noncovalent interactions with MD2 in the MraY AA active site. We applied the quantum theory of atoms in molecules and natural bond orbital analyses based on the density functional theory method on the solved crystal structure of MraY with the MD2 to quantitatively estimate the intermolecular interactions. The obtained results revealed the presence of multiple hydrogen bonds in the investigated active site with strength ranging from van der Waals to covalent limits. Lys70, Asp193, Gly194, Asp196, Gly264, Ala321, Gln305, and His325 are key active-site residues interacting with MD2. Conventional and unconventional hydrogen bonds in addition with charge–dipole and dipole–dipole interactions contribute significantly to stabilize the MD2 binding to the MraY AA active site. It was also found that water molecules inside the active site have substantial effects on its structure stability through hydrogen-bonding interactions with MD2 and the interacting residues.