| 14762003 |
Structural and functional characterization of gene clusters directing nonribosomal synthesis of bioactive cyclic lipopeptides in Bacillus amyloliquefaciens strain FZB42 |
10.1128/JB.186.4.1084-1096.2004. |
J Bacteriol |
Structural and functional characterization of gene clusters directing nonribosomal synthesis of bioactive cyclic lipopeptides in Bacillus amyloliquefaciens strain FZB42
Abstract
- The environmental strain Bacillus amyloliquefaciens FZB42 promotes plant growth and suppresses plant pathogenic organisms present in the rhizosphere. We sampled sequenced the genome of FZB42 and identified 2,947 genes with >50% identity on the amino acid level to the corresponding genes of Bacillus subtilis 168. Six large gene clusters encoding nonribosomal peptide synthetases (NRPS) and polyketide synthases (PKS) occupied 7.5% of the whole genome. Two of the PKS and one of the NRPS encoding gene clusters were unique insertions in the FZB42 genome and are not present in B. subtilis 168. Matrix-assisted laser desorption ionization-time of flight mass spectrometry analysis revealed expression of the antibiotic lipopeptide products surfactin, fengycin, and bacillomycin D. The fengycin (fen) and the surfactin (srf) operons were organized and located as in B. subtilis 168. A large 37.2-kb antibiotic DNA island containing the bmy gene cluster was attributed to the biosynthesis of bacillomycin D. The bmy island was found inserted close to the fen operon. The responsibility of the bmy, fen, and srf gene clusters for the production of the corresponding secondary metabolites was demonstrated by cassette mutagenesis, which led to the loss of the ability to produce these peptides. Although these single mutants still largely retained their ability to control fungal spread, a double mutant lacking both bacillomycin D and fengycin was heavily impaired in its ability to inhibit growth of phytopathogenic fungi, suggesting that both lipopeptides act in a synergistic manner.
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| 14775070 |
[Choline therapy of toxic diseases of the liver] |
10.1007/BF01737300. |
Klin Wochenschr |
[Choline therapy of toxic diseases of the liver]
Abstract
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| 14839088 |
[Identity of the antidiuretic substance of urine in hepatic cirrhosis with adiuretin] |
None |
Boll Soc Ital Biol Sper |
[Identity of the antidiuretic substance of urine in hepatic cirrhosis with adiuretin]
Abstract
|
| 14892017 |
Observations on antiviral activity of viscosin |
10.3181/00379727-78-19071. |
Proc Soc Exp Biol Med |
Observations on antiviral activity of viscosin
Abstract
|
| 14945397 |
Chemical contributions to the mechanism of the biological oxidation of tryptophan |
10.1007/BF02168898. |
Experientia |
Chemical contributions to the mechanism of the biological oxidation of tryptophan
Abstract
|
| 14950216 |
Differentiation between circulins A and B and polymyxins A and E by paper chromatography |
10.1126/science.116.3006.147. |
Science |
Differentiation between circulins A and B and polymyxins A and E by paper chromatography
Abstract
|
| 14962227 |
Prothrombin Shanghai: hypoprothrombinaemia caused by substitution of Gla29 by Gly. |
10.1046/j.1365-2516.2003.00838.x |
Haemophilia |
Prothrombin Shanghai: hypoprothrombinaemia caused by substitution of Gla29 by Gly.
Abstract
- Prothrombin deficiency is a rare bleeding disorder inherited as an autosomal recessive trait. In this study, we reported a Chinese family with hereditary prothrombin deficiency. The proposita had a prolonged activated partial thromboplastin time (APTT, 71.6 s) and prothrombin time (PT, 28.0 s). The coagulation factors activities were normal except that prothrombin coagulation activity was markedly reduced, and the prothrombin antigen level was moderately decreased. Nucleotide sequencing of amplified DNA revealed a novel mutation, Glu (GAG) to Gly (GGG) at residue 29, which normally undergoes gamma-carboxylation within the Gla domain of prothrombin. The proposita was identified as homozygous, while her father, mother and maternal grandmother were heterozygous for the mutation. Gla29 has been demonstrated as one of the key residue for Ca2+-binding, membrane interaction and biological activity of prothrombin.
|
| 14970207 |
Solution structure of the pore-forming protein of Entamoeba histolytica |
10.1074/jbc.M312978200. |
J Biol Chem |
Solution structure of the pore-forming protein of Entamoeba histolytica
Abstract
- Amoebapore A is a 77-residue protein from the protozoan parasite and human pathogen Entamoeba histolytica. Amoebapores lyse both bacteria and eukaryotic cells by pore formation and play a pivotal role in the destruction of host tissues during amoebiasis, one of the most life-threatening parasitic diseases. Amoebapore A belongs to the superfamily of saposin-like proteins that are characterized by a conserved disulfide bond pattern and a fold consisting of five helices. Membrane-permeabilizing effector molecules of mammalian lymphocytes such as porcine NK-lysin and the human granulysin share these structural attributes. Several mechanisms have been proposed to explain how saposin-like proteins form membrane pores. All mechanisms indicate that the surface charge distribution of these proteins is the basis of their membrane binding capacity and pore formation. Here, we have solved the structure of amoebapore A by NMR spectroscopy. We demonstrate that the specific activation step of amoebapore A depends on a pH-dependent dimerization event and is modulated by a surface-exposed histidine residue. Thus, histidine-mediated dimerization is the molecular switch for pore formation and reveals a novel activation mechanism of pore-forming toxins.
|
| 14971903 |
Chemical and functional identification and characterization of novel sulfated alpha-conotoxins from the cone snail Conus anemone |
10.1021/jm031010o. |
J Med Chem |
Chemical and functional identification and characterization of novel sulfated alpha-conotoxins from the cone snail Conus anemone
Abstract
- An LC/MS analysis with diagnostic screening for the detection of peptides with posttranslational modifications revealed the presence of novel sulfated peptides within the alpha-conotoxin molecular mass range in Conus anemone crude venom. A functional assay of the extract showed activity at several neuronal nicotinic acetylcholine receptors (nAChRs). Three sulfated alpha-conotoxins (AnIA, AnIB, and AnIC) were identified by LC/MS and assay-directed fractionation and sequenced after purification. The most active of these, alpha-AnIB, was further characterized and used to investigate the influence of posttranslational modifications on affinity. Synthetic AnIB exhibited subnanomolar potency at the rat alpha3beta2 nAChR (IC50 0.3 nM) and was 200-fold less active on the rat alpha7 nAChR (IC50 76 nM). The unsulfated peptide [Tyr16]AnIB showed a 2-fold and 10-fold decrease in activities at alpha3beta2 (IC50 0.6 nM) and alpha7 (IC50 836 nM) nAChR, respectively. Likewise, removal of the C-terminal amide had a greater influence on potency at the alpha7 (IC50 367 nM) than at the alpha3beta2 nAChR (IC50 0.5 nM). Stepwise removal of two N-terminal glycine residues revealed that these residues affect the binding kinetics of the peptide. Comparison with similar 4/7-alpha-conotoxin sequences suggests that residue 11 (alanine or glycine) and residue 14 (glutamine) constitute important determinants for alpha3beta2 selectivity, whereas the C-terminal amidation and sulfation at tyrosine-16 favor alpha7 affinity.
|
| 14978308 |
Lead optimization of antifungal peptides with 3D NMR structures analysis |
10.1110/ps.03404404. |
Protein Sci |
Lead optimization of antifungal peptides with 3D NMR structures analysis
Abstract
- Antimicrobial peptides are key components of the innate immune response in most multicellular organisms. These molecules are considered as one of the most innovative class of anti-infective agents that have been discovered over the last two decades, and therefore, as a source of inspiration for novel drug design. Insect cystine-rich antimicrobial peptides with the CS alpha beta scaffold (an alpha-helix linked to a beta-sheet by two disulfide bridges) represent particularly attractive templates for the development of systemic agents owing to their remarkable resistance to protease degradation. We have selected heliomicin, a broad spectrum antifungal CS alpha beta peptide from Lepidoptera as the starting point of a lead optimization program based on phylogenic exploration and fine tuned mutagenesis. We report here the characterization, biological activity, and 3D structure of heliomicin improved analogs, namely the peptides ARD1, ETD-135, and ETD-151. The ARD1 peptide was initially purified from the immune hemolymph of the caterpillars of Archeoprepona demophoon. Although it differs from heliomicin by only two residues, it was found to be more active against the human pathogens Aspergillus fumigatus and Candida albicans. The peptides ETD-135 and ETD-151 were engineered by site-directed mutagenesis of ARD1 in either cationic or hydrophobic regions. ETD-135 and ETD-151 demonstrated an improved antifungal activity over the native peptides, heliomicin and ARD1. A comparative analysis of the 3D structure of the four molecules highlighted the direct impact of the modification of the amphipathic properties on the molecule potency. In addition, it allowed to characterize an optimal organization of cationic and hydrophobic regions to achieve best antifungal activity.
|
| 14987049 |
Cytotoxic cyclotides from Viola tricolor |
10.1021/np030101l. |
J Nat Prod |
Cytotoxic cyclotides from Viola tricolor
Abstract
- A crude fraction of Viola tricolor rich in small lipophilic proteins was prepared and subjected to fractionation guided by bioactivity, using RP-HPLC and a fluorometric cytotoxicity assay. Two human cancer cell lines, U-937 GTB (lymphoma) and RPMI-8226/s (myeloma), were used in this study. The most potent compounds isolated, that is, the compounds showing the lowest IC(50) values, were shown to be three small proteins: vitri A (IC(50) = 0.6 microM and IC(50) = 1 microM, respectively), varv A (IC(50) = 6 microM and IC(50) = 3 microM, respectively), and varv E (IC(50) = 4 microM in both cell lines). Their sequences, determined by automated Edman degradation, quantitative amino acid analysis, and mass spectrometry, were cyclo-GESCVWIPCITSAIGCSCKSKVCYRNGIPC (vitri A), cyclo-GETCVGGTCNTPGCSCSWPVCTRNGLPVC (varv A), and cyclo-GETCVGGTCNTPGCSCSWPVCTRNGLPIC (varv E), of which vitri A is described for the first time. Each forms a head-to-tail cyclic backbone, with six cysteine residues being involved in three disulfide bonds, characteristic of the family of small proteins called the cyclotides. This is the first report on cyclotides from the species V. tricolor and the first report on the sequence of the cytotoxic cyclotide vitri A.
|
| 14996845 |
Identification and characterization of a mucosal antimicrobial peptide expressed by the chinchilla (Chinchilla lanigera) airway |
10.1074/jbc.M400499200. |
J Biol Chem |
Identification and characterization of a mucosal antimicrobial peptide expressed by the chinchilla (Chinchilla lanigera) airway
Abstract
- Cationic antimicrobial peptides (APs) are produced at mucosal surfaces and play a key role as a first line of defense against infection. To understand how APs might impact disease progression in otitis media (OM), our goal was to identify and characterize APs expressed by the epithelium lining the uppermost airway of the chinchilla, the established rodent host for the study of the bacterial-viral pathogenesis in OM. Using a molecular approach, we cloned a cDNA encoding a homolog of human beta-defensin 3, designated chinchilla beta-defensin-1 (cBD-1), and found by Northern analysis expression of the corresponding mRNA in nasopharyngeal and tongue mucosae as well as skin. By reverse transcription-PCR, cBD-1 mRNA was also detected in RNA isolated from trachea, lung, and Eustachian tube tissues. The predicted mature form of cBD-1, expressed as a recombinant peptide in Escherichia coli, demonstrated bactericidal activity against the three primary opportunistic pathogens of OM as well as Candida albicans. Continued study of this and other APs will allow us to determine their role in bacterial colonization of the upper airway as well as how viruses might contribute to the pathogenesis of OM by modulating AP expression.
|
| 15003829 |
A family of brevinin-2 peptides with potent activity against Pseudomonas aeruginosa from the skin of the Hokkaido frog, Rana pirica |
10.1016/j.regpep.2003.12.003. |
Regul Pept |
A family of brevinin-2 peptides with potent activity against Pseudomonas aeruginosa from the skin of the Hokkaido frog, Rana pirica
Abstract
- Nine peptides displaying varying degrees of antimicrobial activity were extracted from the skin of the Hokkaido frog, Rana pirica. Five structurally related peptides were identified as members of the brevinin-2 family. These peptides were active against reference strains of Gram-negative (Escherichia coli, Pseudomonas aeruginosa, Enterobacter cloacae, Klebsiella pneumoniae) and Gram-positive (Staphlococcus aureus) bacteria but displayed relatively low hemolytic activity. The most abundant peptide, brevinin-2PRa (680 nmol/g weight of dry skin) showed high potency [minimal inhibitory concentration (MIC) values between 6 and 12 microM] against a range of clinical isolates of P. aeruginosa. In addition, activity was unaffected by NaCl concentrations up to 200 mM. Cladistic analysis based on the primary structures of brevinin-2 peptides supports a close phylogenetic relationship between R. pirica and Japanese mountain brown frog Rana ornativentris. One peptide of the ranatuerin-2 family and one strongly hemolytic peptide of the brevinin-1 family were also isolated from the extract along with two members of the temporin family, temporin-1PRa (ILPILGNLLNGLL.NH(2)) and temporin-1PRb (ILPILGNLLNSLL.NH(2)) that atypically lacked basic amino acid residues and showed only very weak antimicrobial and hemolytic activity.
|
| 15005608 |
Determinants of potency on alpha-conotoxin MII, a peptide antagonist of neuronal nicotinic receptors |
10.1021/bi036180h. |
Biochemistry |
Determinants of potency on alpha-conotoxin MII, a peptide antagonist of neuronal nicotinic receptors
Abstract
- Alpha-conotoxin MII, a peptide toxin isolated from Conus magus, antagonizes a subset of neuronal nicotinic receptors. Rat alpha3beta2 receptors, expressed in Xenopus oocytes, are blocked with an IC(50) of 3.7 +/- 0.3 nM. To identify structural features that determine toxin potency, a series of alanine-substituted toxins were synthesized and tested for the ability to block the function of alpha3beta2 receptors. Circular dichroism and protein modeling were used to assess the structural integrity of the mutant toxins. Three residues were identified as major determinants of toxin potency. Replacement of asparagine 5, proline 6, or histidine 12 with alanine resulted in >2700-fold, 700-fold, and approximately 2700-fold losses in toxin potency, respectively. A decrease in pH improved toxin potency, while an increase in pH eliminated toxin blockade, suggesting that, in the active form of the toxin, histidine 12 is charged. The imidazole ring of histidine 12 protrudes from one side, while asparagine 5 and proline 6 are located at the opposite end of the toxin structure. The side chains of these three residues are exposed on the surface of the toxin, suggesting that they directly interact with the alpha3beta2 receptor.
|
| 15023056 |
Structural characterization of lacticin 3147, a two-peptide lantibiotic with synergistic activity |
10.1021/bi0362065. |
Biochemistry |
Structural characterization of lacticin 3147, a two-peptide lantibiotic with synergistic activity
Abstract
- Lantibiotics are antibacterial peptides isolated from bacterial sources that exhibit activity toward Gram-positive organisms and are usually several orders of magnitude more potent than traditional antibiotics such as penicillin. They contain a number of unique structural features including dehydro amino acid and lanthionine (thioether) residues. Introduced following ribosomal translation of the parent peptide, these moieties render conventional methods of peptide analysis ineffective. We report herein a new method using nickel boride (Ni(2)B), in the presence of deuterium gas, to reduce dehydro side chains and reductively desulfurize lanthionine bridges found in lantibiotics. Using this approach, it is possible to identify and distinguish the original locations of dehydro side chains and lanthionine bridges by traditional peptide sequencing (Edman degradation) followed by mass spectrometry. The strategy was initially verified using nisin A, a structurally well characterized lantibiotic, and subsequently extended to the novel two-component lantibiotic, lacticin 3147, produced by Lactococcus lactis subspecies lactis DPC3147. The primary structures of both lacticin 3147 peptides were then fully assigned by use of multidimensional NMR spectroscopy, showing that lacticin 3147 A1 has a specific lanthionine bridging pattern which resembles the globular type-B lantibiotic mersacidin, whereas the A2 peptide is a member of the elongated type-A lantibiotic class. Also obtained by NMR were solution conformations of both lacticin 3147 peptides, indicating that A1 may adopt a conformation similar to that of mersacidin and that the A2 peptide adopts alpha-helical structure. These results are the first of their kind for a synergistic lantibiotic pair (only four such pairs have been reported to date).
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