Structure-function analysis of tritrypticin, an antibacterial peptide of innate immune origin.

The structural requirements for the antibacterial activity of a pseudosymmetric 13-residue peptide, tritrypticin, were analyzed by combining pattern recognition in protein structures, the structure-activity knowledge-base, and circular dichroism. The structure-activity analysis, based on various deletion analogs, led to the identification of two minimal functional peptides, which by themselves exhibit adequate antibacterial activity against Escherichia coli and Salmonella typhimurium. The common features between these two peptides are that they both share an aromatic-proline-aromatic (ArProAr) sequence motif, and their sequences are retro with respect to one another. The pattern searches in protein structure data base using the ArProAr motif led to the identification of two distinct conformational clusters, namely polyproline type II and beta-turn, which correspond to the observed solution structures of the two minimal functional analogs. The role of different residues in structure and function of tritrypticin was delineated by analyzing antibacterial activity and circular dichroism spectra of various designed analogs. Three main results arise from this study. First, the ArProAr sequence motif in proteins has definitive conformational features associated with it. Second, the two minimal bioactive domains of tritrypticin have entirely different structures while having equivalent activities. Third, tritrypticin has a beta-turn conformation in solution, but the functionally relevant conformation of this gene-encoded peptide antibiotic may be an extended polyproline type II.     

Screening of phytotoxicity of Alternaria macrospora MKP1 against Parthenium hysterophorus L.

Parthenium hysterophorus L. an exotic, pernicious weed is considered as one of the most troublesome weeds for agricultural sector by virtue of its high ecological amplitude and adaptability. Microbes and their by-products are now proved to be a worthy alternative to toxic chemicals used for weed management. Alternaria macrospora MKPI was isolated from the parthenium leaves infected with leaf blight and found pathogenic to the weed. The herbicidal potential of cell free culture filtrate of A. macrospora MKP1 has been tested against parthenium by employing detached leaf bioassay and seed germination bioassay and a significant damage was exhibited by the cultural filtrate of pathogen to the parthenium leaves and seeds.     

CUX2 Protein Functions as an Accessory Factor in the Repair of Oxidative DNA Damage

CUX1 and CUX2 proteins are characterized by the presence of three highly similar regions called Cut repeats 1, 2, and 3. Although CUX1 is ubiquitously expressed, CUX2 plays an important role in the specification of neuronal cells and continues to be expressed in postmitotic neurons. Cut repeats from the CUX1 protein were recently shown to stimulate 8-oxoguanine DNA glycosylase 1 (OGG1), an enzyme that removes oxidized purines from DNA and introduces a single strand break through its apurinic/apyrimidinic lyase activity to initiate base excision repair. Here, we investigated whether CUX2 plays a similar role in the repair of oxidative DNA damage. Cux2 knockdown in embryonic cortical neurons increased levels of oxidative DNA damage. In vitro, Cut repeats from CUX2 increased the binding of OGG1 to 7,8-dihydro-8-oxoguanine-containing DNA and stimulated both the glycosylase and apurinic/apyrimidinic lyase activities of OGG1. Genetic inactivation in mouse embryo fibroblasts or CUX2 knockdown in HCC38 cells delayed DNA repair and increased DNA damage. Conversely, ectopic expression of Cut repeats from CUX2 accelerated DNA repair and reduced levels of oxidative DNA damage. These results demonstrate that CUX2 functions as an accessory factor that stimulates the repair of oxidative DNA damage. Neurons produce a high level of reactive oxygen species because of their dependence on aerobic oxidation of glucose as their source of energy. Our results suggest that the persistent expression of CUX2 in postmitotic neurons contributes to the maintenance of genome integrity through its stimulation of oxidative DNA damage repair.     

Long-term stability in biomass and production of terpene indole alkaloids by hairy root culture of Rauvolfia serpentina and cost approximation to endorse commercial realism

The effect of 6 years of cultivation and use of table-sugar (TS) on the biomass/terpene alkaloid productivities and rol gene expression were studied in a hairy root (HR) clone of Rauvolfia serpentina. The media cost could be reduced >94 % by replacing sucrose (SUC) with TS—an unexplored avenue for HR cultivation. The overall productivities increased over long-term cultivation with sugar proving superior to SUC for biomass (24.4 ± 2.11 g/l DW after 40 days to 17.31 % higher) and reserpine (0.094 ± 0.008 % DW after 60 days to 193.8 % more) production. The latter however revealed comparatively better yields concerning ajmaline (0.507 ± 0.048 % DW after 60 days to 61.98 % higher) and yohimbine (0.628 ± 0.062 % DW after 60 days to 38.32 % higher), respectively. PCR amplification of rol genes confirmed long-term expression stability.     

Fluorescent labeling of tRNA dihydrouridine residues: Mechanism and distribution

Dihydrouridine (DHU) positions within tRNAs have long been used as sites to covalently attach fluorophores, by virtue of their unique chemical reactivity toward reduction by NaBH(4), their abundance within prokaryotic and eukaryotic tRNAs, and the biochemical functionality of the labeled tRNAs so produced. Interpretation of experiments employing labeled tRNAs can depend on knowing the distribution of dye among the DHU positions present in a labeled tRNA. Here we combine matrix-assisted laser desorption/ionization mass spectroscopy (MALDI-MS) analysis of oligonucleotide fragments and thin layer chromatography to resolve and quantify sites of DHU labeling by the fluorophores Cy3, Cy5, and proflavin in Escherichia coli tRNA(Phe) and E. coli tRNA(Arg). The MALDI-MS results led us to re-examine the precise chemistry of the reactions that result in fluorophore introduction into tRNA. We demonstrate that, in contrast to an earlier suggestion that has long been unchallenged in the literature, such introduction proceeds via a substitution reaction on tetrahydrouridine, the product of NaBH(4) reduction of DHU, resulting in formation of substituted tetrahydrocytidines within tRNA.     

DDESC: Dragon database for exploration of sodium channels in human

Background

Hepcidin/LEAP-1 is an iron regulatory hormone originally identified as an antimicrobial peptide. As part of a systematic analysis of the evolution of host defense peptides in primates, we have sequenced the orthologous gene from 14 species of non-human primates.

Results

The sequence of the mature peptide is highly conserved amongst all the analyzed species, being identical to the human one in great apes and gibbons, with a single residue conservative variation in Old-World monkeys and with few substitutions in New-World monkeys.

Conclusion

Our analysis indicates that hepcidin's role as a regulatory hormone, which involves interaction with a conserved receptor (ferroportin), may result in conservation over most of its sequence, with the exception of the stretch between residues 15 and 18, which in New-World monkeys (as well as in other mammals) shows a significant variation, possibly indicating that this structural region is involved in other functions.     

Cj1121c, a novel UDP-4-keto-6-deoxy-GlcNAc C-4 aminotransferase essential for protein glycosylation and virulence in Campylobacter jejuni

Campylobacter jejuni produces glycoproteins that are essential for virulence. These glycoproteins carry diacetamidobacillosamine (DAB), a sugar that is not found in humans. Hence, the enzymes responsible for DAB synthesis represent potential therapeutic targets. We describe the biochemical characterization of Cj1121c, a putative aminotransferase encoded by the general protein glycosylation locus, to assess its role in DAB biosynthesis. By using overexpressed and affinity-purified enzyme, we demonstrate that Cj1121c has pyridoxal phosphate- and glutamate-dependent UDP-4-keto-6-deoxy-GlcNAc C-4 transaminase activity and produces UDP-4-amino-4,6-dideoxy-GlcNAc. This is consistent with a role in DAB biosynthesis and distinguishes Cj1121c from Cj1294, a homologous UDP-2-acetamido-2,6-dideoxy-beta-l-arabino-4-hexulose C-4 aminotransferase that we characterized previously. We show that Cj1121c can also use this 4-keto-arabino sugar indirectly as a substrate, that Cj1121c and Cj1294 are active simultaneously in C. jejuni, and that the activity of Cj1121c is preponderant under standard growth conditions. Kinetic data indicate that Cj1121c has a slightly higher catalytic efficiency than Cj1294 with regard to the 4-keto-arabino substrate. By site-directed mutagenesis, we show that residues Glu-158 and Leu-131 are not essential for catalysis or for substrate specificity contrary to expectations. We further demonstrate that a cj1121c knock-out mutant is impaired for flagella-mediated motility, for invasion of intestinal epithelial cells, and for persistence in the chicken intestine, clearly demonstrating that Cj1121c is essential for host colonization and virulence. Finally, we show that cj1121c is necessary for protein glycosylation by lectin Western blotting. Collectively, these results validate Cj1121c as a promising drug target and provide the means to assay for inhibitors.     

Opioid Peptides: An Overview of Functional Significance

International Journal of Peptide Research and Therapeutics - Bioactive peptides have been reported to exhibit opioid-like activity and are termed as opioid peptides. Either these are produced...