| 16755625 |
The fumitremorgin gene cluster of Aspergillus fumigatus: identification of a gene encoding brevianamide F synthetase |
10.1002/cbic.200600003. |
Chembiochem |
The fumitremorgin gene cluster of Aspergillus fumigatus: identification of a gene encoding brevianamide F synthetase
Abstract
- A gene encoding a putative dimodular nonribosomal peptide synthetase (NRPS) was identified within a gene cluster of Aspergillus fumigatus, a species reported to produce fumitremorgins and other prenylated alkaloids. The gene was deleted and overexpressed in the genome reference strain Af293, and was also expressed in the naïve host Aspergillus nidulans, which lacks the equivalent gene cluster. While neither fumitremorgins nor the dipeptide brevianamide F (cyclo-L-Trp-L-Pro), an early intermediate, were detected in wild-type and deletion strains of A. fumigatus, brevianamide F accumulated in fungal cultures following increased expression of the NRPS gene in both A. fumigatus and A. nidulans. We conclude that the gene Afu8g00170, named ftmA, encodes the NRPS brevianamide synthetase. Brevianamide F is the precursor of a variety of fungal prenylated alkaloids with biological activity, including fumitremorgins A, B and C and tryprostatin B.
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| 16756302 |
Lariatins, antimycobacterial peptides produced by Rhodococcus sp. K01-B0171, have a lasso structure |
10.1021/ja056780z. |
J Am Chem Soc |
Lariatins, antimycobacterial peptides produced by Rhodococcus sp. K01-B0171, have a lasso structure
Abstract
- Two antimycobacterial agents, lariatins A and B, were isolated from the culture broth of Rhodococcus sp. K01-B0171. Their structures were elucidated by spectral analysis and advanced protein chemical methods to be unique cyclic peptides, which consist of 18 and 20 L-amino acid residues with an internal linkage between the gamma-carboxyl group of Glu8 and the alpha-amino group of Gly1. The three-dimensional structure of lariatin A deduced from NMR data by dynamical simulated annealing method indicates that the tail segment (Trp9-Pro18) passes through the ring segment (Gly1-Glu8) to form a 'lasso' structure.
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| 16763681 |
Multipolar interactions in the D pocket of thrombin: large differences between tricyclic imide and lactam inhibitors |
10.1039/b602585d. |
Org Biomol Chem |
Multipolar interactions in the D pocket of thrombin: large differences between tricyclic imide and lactam inhibitors
Abstract
- Two series of tricyclic inhibitors of the serine protease thrombin, imides (+/-)-1-(+/-)-8 and lactams (+/-)-9-(+/-)-13, were analysed to evaluate contributions of orthogonal multipolar interactions with the backbone C=O moiety of Asn98 to the free enthalpy of protein-ligand complexation. The lactam derivatives are much more potent and more selective inhibitors (K(i) values between 0.065 and 0.005 microM, selectivity for thrombin over trypsin between 361- and 1609-fold) than the imide compounds (Ki values between 0.057 and 23.7 microM, selectivity for thrombin over trypsin between 3- and 67-fold). The increase in potency and selectivity is explained by the favorable occupancy of the P-pocket of thrombin by the additional isopropyl substituent in the lactam derivatives. The nature of the substituent on the benzyl ring filling the D pocket strongly influences binding potency in the imide series, with Ki values increasing in the sequence: F < OCH2O < Cl < H < OMe < OH < N(pyr)<< Br. This sequence can be explained by both steric fit and the occurrence of orthogonal multipolar interactions with the backbone C[double bond, length as m-dash]O moiety of Asn98. In contrast, the substituent on the benzyl ring hardly affects the ligand potency in the lactam series. This discrepancy was clarified by the comparison of X-ray structures solved for co-crystals of thrombin with imide and lactam ligands. Whereas the benzyl substituents in the imide inhibitors are sufficiently close (< or =3.5 Angstroms) to the C=O group of Asn98 to allow for attractive orthogonal multipolar interactions, the distances in the lactam series are too large (> or =4 Angstroms) for attractive dipolar contacts to be effective.
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| 16765485 |
Direct cDNA cloning of novel conotoxins of the T-superfamily from Conus textile |
10.1016/j.peptides.2006.05.002. |
Peptides |
Direct cDNA cloning of novel conotoxins of the T-superfamily from Conus textile
Abstract
- The T-superfamily is a large and diverse group of peptides, widely distributed in venom ducts of all major feeding types of Conus. These peptides are likely to be functionally diverse. A directed PCR-based approach using primers based on the conserved signal sequence was applied to investigate new conotoxins of the T-superfamily from Conus textile native to Hainan. Using RT-PCR and 3'-RACE, four novel cDNA sequences encoding precursor peptides were identified in C. textile. They share a common T-superfamily cysteine pattern (CC-CC, with two disulfide bridges). The predicted peptides are small (9-12 amino acids). TeAr193 composed of nine amino acid residues is one of the shortest T-superfamily conotoxins ever found. Patterns of sequence divergence and Cys codon usage define the major T-superfamily branches and suggest how these separate branches arose. The sequences of the signal regions exhibited highest conservation, whereas the sequences of the mature peptides were either almost identical or highly divergent; and conservation of the pro-region was intermediate between that observed in signal and toxin regions. The elucidated cDNAs of the four toxins will facilitate a better understanding of the relationship between structure and function.
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| 16766795 |
The absolute structural requirement for a proline in the P3'-position of Bowman-Birk protease inhibitors is surmounted in the minimized SFTI-1 scaffold |
10.1074/jbc.M601426200. |
J Biol Chem |
The absolute structural requirement for a proline in the P3'-position of Bowman-Birk protease inhibitors is surmounted in the minimized SFTI-1 scaffold
Abstract
- SFTI-1 is a small cyclic peptide from sunflower seeds that is one of the most potent trypsin inhibitors of any naturally occurring peptide and is related to the Bowman-Birk family of inhibitors (BBIs). BBIs are involved in the defense mechanisms of plants and also have potential as cancer chemopreventive agents. At only 14 amino acids in size, SFTI-1 is thought to be a highly optimized scaffold of the BBI active site region, and thus it is of interest to examine its important structural and functional features. In this study, a suite of 12 alanine mutants of SFTI-1 has been synthesized, and their structures and activities have been determined. SFTI-1 incorporates a binding loop that is clasped together with a disulfide bond and a secondary peptide loop making up the circular backbone. We show here that the secondary loop stabilizes the binding loop to the consequences of sequence variations. In particular, full-length BBIs have a conserved cis-proline that has been shown previously to be required for well defined structure and potent activity, but we show here that the SFTI-1 scaffold can accommodate mutation of this residue and still have a well defined native-like conformation and nanomolar activity in inhibiting trypsin. Among the Ala mutants, the most significant structural perturbation occurred when Asp14 was mutated, and it appears that this residue is important in stabilizing the trans peptide bond preceding Pro13 and is thus a key residue in maintaining the highly constrained structure of SFTI-1. This aspartic acid residue is thought to be involved in the cyclization mechanism associated with excision of SFTI-1 from its 58-amino acid precursor. Overall, this mutational analysis of SFTI-1 clearly defines the optimized nature of the SFTI-1 scaffold and demonstrates the importance of the secondary loop in maintaining the active conformation of the binding loop.
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| 16779750 |
Anidulafungin: a novel echinocandin |
10.1086/505204. |
Clin Infect Dis |
Anidulafungin: a novel echinocandin
Abstract
- Until recently, the treatment available for serious fungal infections was composed of amphotericin B and azoles, and each class demonstrated significant limitations. Echinocandins are a new class of drugs that have shown promising results in treating a variety of fungal infections. Of these, anidulafungin is a novel echinocandin that appears to have several advantages over existing antifungals. It is unique because it slowly degrades in humans, undergoing a process of biotransformation rather than being metabolized. It has potent in vitro activity against Aspergillus and Candida species, including those resistant to fluconazole or amphotericin B. Results of several clinical trials indicate that anidulafungin is effective in treating esophageal candidiasis, including azole-refractory disease. The results of a recent study comparing fluconazole versus anidulafungin demonstrated the superiority of anidulafungin in the treatment of candidemia and invasive candidiasis (IC). Studies evaluating the concomitant use of anidulafungin and either amphotericin B, voriconazole, or cyclosporine did not demonstrate significant drug-drug interactions or adverse events. To date, anidulafungin appears to have an excellent safety profile. On the basis of early clinical experience, it appears that anidulafungin will be a valuable asset in the management of serious and difficult-to-treat fungal infections.
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| 16781016 |
Antifouling activity of synthesized peptide analogs of the sponge metabolite barettin |
10.1016/j.peptides.2006.03.027. |
Peptides |
Antifouling activity of synthesized peptide analogs of the sponge metabolite barettin
Abstract
- Barettin (cyclo [(6-bromo-8-en-tryptophan) arginine]), a diketopiperazine isolated from the marine sponge Geodia barretti, is a potent inhibitor of barnacle larvae settlement with an EC50-value of 0.9 microM. In the present study, 14 analogs of barettin and its structural congener dipodazine were synthezised and tested for their ability to inhibit larval settlement. Two of the analogs have an intact barettin skeleton. The remaining analogs have a dipodazine skeleton (a diketopiperazine where arginine is replaced with glycine). Six of the tested synthetic analogs displayed significant settlement inhibition with the most potent inhibitor being benzo[g]dipodazine, which displayed even stronger activity than barettin (EC50-value 0.034 microM). The effect of benzo[g]dipodazine was also shown to be readily reversible, when cyprids were transferred to filtered seawater (FSW).
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| 16785532 |
The biological activity of human CD20 monoclonal antibodies is linked to unique epitopes on CD20 |
10.4049/jimmunol.177.1.362. |
J Immunol |
The biological activity of human CD20 monoclonal antibodies is linked to unique epitopes on CD20
Abstract
- We have previously defined a panel of fully human CD20 mAb. Most of these were unexpectedly efficient in their ability to recruit C1q to the surface of CD20-positive cells and mediate tumor lysis via activation of the classical pathway of complement. This complement-dependent cytotoxicity (CDC) potency appeared to relate to the unusually slow off-rate of these human Abs. However, we now present epitope-mapping data, which indicates that all human mAb bind a novel region of CD20 that may influence CDC potency. Epitope mapping, using both mutagenesis studies and overlapping 15-mer peptides of the extracellular loops of CD20, defined the amino acids required for binding by an extensive panel of mouse and human mAb. Binding by rituximab and mouse CD20 mAb, had an absolute requirement for alanine and proline at positions 170 and 172, respectively, within the large extracellular loop of CD20. Surprisingly, however, all of the human CD20 mAb recognize a completely novel epitope located N-terminally of this motif, also including the small extracellular loop of CD20. Thus, although off-rate may influence biological activity of mAb, another critical factor for determining CDC potency by CD20 mAb appears to be the region of the target molecule they recognize. We conclude that recognition of the novel epitope cooperates with slow off-rate in determining the activity of CD20 Ab in activation of complement and induction of tumor cell lysis.
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| 16790424 |
Identification of a novel class of nicotinic receptor antagonists: dimeric conotoxins VxXIIA, VxXIIB, and VxXIIC from Conus vexillum |
10.1074/jbc.M603703200. |
J Biol Chem |
Identification of a novel class of nicotinic receptor antagonists: dimeric conotoxins VxXIIA, VxXIIB, and VxXIIC from Conus vexillum
Abstract
- The venoms of predatory marine snails (Conus spp.) contain diverse mixtures of peptide toxins with high potency and selectivity for a variety of voltage-gated and ligand-gated ion channels. Here we describe the chemical and functional characterization of three novel conotoxins, alphaD-VxXIIA, alphaD-VxXIIB, and alphaD-VxXIIC, purified from the venom of Conus vexillum. Each toxin was observed as an approximately 11-kDa protein by LC/MS, size exclusion chromatography, and SDS-PAGE. After reduction, the peptide sequences were determined by Edman degradation chemistry and tandem MS. Combining the sequence data together with LC/MS and NMR data revealed that in solution these toxins are pseudo-homodimers of paired 47-50-residue peptides. The toxin subunits exhibited a novel arrangement of 10 conserved cystine residues, and additional post-translational modifications contributed heterogeneity to the proteins. Binding assays and two-electrode voltage clamp analyses showed that alphaD-VxXIIA, alphaD-VxXIIB, and alphaD-VxXIIC are potent inhibitors of nicotinic acetylcholine receptors (nAChRs) with selectivity for alpha7 and beta2 containing neuronal nAChR subtypes. These dimeric conotoxins represent a fifth and highly divergent structural class of conotoxins targeting nAChRs.
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| 16803542 |
Results with EMA/CO (etoposide, methotrexate, actinomycin D, cyclophosphamide, vincristine) chemotherapy in gestational trophoblastic neoplasia |
10.1111/j.1525-1438.2006.00606.x. |
Int J Gynecol Cancer |
Results with EMA/CO (etoposide, methotrexate, actinomycin D, cyclophosphamide, vincristine) chemotherapy in gestational trophoblastic neoplasia
Abstract
- The aim of this study was to evaluate the efficacy and toxicity of EMA/CO (etoposide, methotrexate, actinomycin D, cyclophosphamide, vincristine) regimen for the treatment of high-risk gestational trophoblastic neoplasia (GTN). Thirty-three patients with high-risk GTN, scored according to World Health Organization, received 159 EMA/CO treatment cycles between 1994 and 2004. Twenty-three patients were treated primarily with EMA/CO, and 10 patients were treated secondarily after failure of single agent or MAC (methotrexate, actinomycin D, cyclophosphamide, or clorambucile) III chemotherapy. Adjuvant surgery and radiotherapy were used in selected patients. Survival, response, and toxicity were analyzed retrospectively. The overall survival rate was 90.9% (30/33). Survival rates were 91.3% (21/23) for primary treatment and 90% (9/10) for secondary treatment. Six (18.2%) of 33 patients had drug resistance. Four of them underwent surgery for adjuvant therapy. Three of these patients with drug resistance died. Survival and complete response to EMA/CO were influenced by liver metastasis, antecedent pregnancy, and histopathologic diagnosis of choriocarcinoma. Survival rate was also affected by blood group. The treatment was well tolerated. The most severe toxicity was grade 3-4 leukopenia that occurred in 24.3% (8/33) of patients and 6.9% (11/159) of treatment cycles. Febrile neutropenia occurred in one patient (3%). EMA/CO regimen is highly effective for treatment of high-risk GTN. Its toxicity is well tolerated.
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| 16817894 |
Conformation and mode of membrane interaction in cyclotides. Spatial structure of kalata B1 bound to a dodecylphosphocholine micelle |
10.1111/j.1742-4658.2006.05282.x. |
FEBS J |
Conformation and mode of membrane interaction in cyclotides. Spatial structure of kalata B1 bound to a dodecylphosphocholine micelle
Abstract
- Cyclotides are a family of bioactive plant peptides that are characterized by a circular protein backbone and three conserved tightly packed disulfide bonds. The antimicrobial and hemolytic properties of cyclotides, along with the relative hydrophobicity of the peptides, point to the biological membrane as a target for cyclotides. To assess the membrane-induced conformation and orientation of cyclotides, the interaction of the Möbius cyclotide, kalata B1, from the African perennial plant Oldenlandia affinis, with dodecylphosphocholine micelles was studied using NMR spectroscopy. Under conditions where the cyclotide formed a well-defined complex with micelles, the spatial structure of kalata B1 was calculated from NOE and J couplings data, and the model for the peptide-micelle complex was built using 5- and 16-doxylstearate relaxation probes. The binding of divalent cations to the peptide-micelle complex was quantified by Mn2+ titration. The results show that the peptide binds to the micelle surface, with relatively high affinity, via two hydrophobic loops (loop 5, Trp19-Val21; and loop6, Leu27-Val29). The charged residues (Glu3 and Arg24), along with the cation-binding site (near Glu3) are segregated on the other side of the molecule and in contact with polar head groups of detergent. The spatial structure of kalata B1 is only slightly changed during incorporation into micelles and represents a distorted triple-stranded beta-sheet cross-linked by a cystine knot. Detailed structural analysis and comparison with other knottins revealed structural conservation of the two-disulfide motif in cyclic and acyclic peptides. The results thus obtained provide the first model for interaction of cyclotides with membranes and permit consideration of the cyclotides as membrane-active cationic antimicrobial peptides.
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| 16819832 |
A novel conotoxin inhibitor of Kv1.6 channel and nAChR subtypes defines a new superfamily of conotoxins |
10.1021/bi060263r. |
Biochemistry |
A novel conotoxin inhibitor of Kv1.6 channel and nAChR subtypes defines a new superfamily of conotoxins
Abstract
- Using assay-directed fractionation of the venom from the vermivorous cone snail Conus planorbis, we isolated a new conotoxin, designated pl14a, with potent activity at both nicotinic acetylcholine receptors and a voltage-gated potassium channel subtype. pl14a contains 25 amino acid residues with an amidated C-terminus, an elongated N-terminal tail (six residues), and two disulfide bonds (1-3, 2-4 connectivity) in a novel framework distinct from other conotoxins. The peptide was chemically synthesized, and its three-dimensional structure was demonstrated to be well-defined, with an alpha-helix and two 3(10)-helices present. Analysis of a cDNA clone encoding the prepropeptide precursor of pl14a revealed a novel signal sequence, indicating that pl14a belongs to a new gene superfamily, the J-conotoxin superfamily. Five additional peptides in the J-superfamily were identified. Intracranial injection of pl14a in mice elicited excitatory symptoms that included shaking, rapid circling, barrel rolling, and seizures. Using the oocyte heterologous expression system, pl14a was shown to inhibit both a K+ channel subtype (Kv1.6, IC50 = 1.59 microM) and neuronal (IC50 = 8.7 microM for alpha3beta4) and neuromuscular (IC50 = 0.54 microM for alpha1beta1 epsilondelta) subtypes of the nicotinic acetylcholine receptor (nAChR). Similarities in sequence and structure are apparent between the middle loop of pl14a and the second loop of a number of alpha-conotoxins. This is the first conotoxin shown to affect the activity of both voltage-gated and ligand-gated ion channels.
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| 16820529 |
SAP30L interacts with members of the Sin3A corepressor complex and targets Sin3A to the nucleolus |
10.1093/nar/gkl401. |
Nucleic Acids Res |
SAP30L interacts with members of the Sin3A corepressor complex and targets Sin3A to the nucleolus
Abstract
- Histone acetylation plays a key role in the regulation of gene expression. The chromatin structure and accessibility of genes to transcription factors is regulated by enzymes that acetylate and deacetylate histones. The Sin3A corepressor complex recruits histone deacetylases and in many cases represses transcription. Here, we report that SAP30L, a close homolog of Sin3-associated protein 30 (SAP30), interacts with several components of the Sin3A corepressor complex. We show that it binds to the PAH3/HID (Paired Amphipathic Helix 3/Histone deacetylase Interacting Domain) region of mouse Sin3A with residues 120-140 in the C-terminal part of the protein. We provide evidence that SAP30L induces transcriptional repression, possibly via recruitment of Sin3A and histone deacetylases. Finally, we characterize a functional nucleolar localization signal in SAP30L and show that SAP30L and SAP30 are able to target Sin3A to the nucleolus.
|
| 16824043 |
Identification of a cowpea gamma-thionin with bactericidal activity |
10.1111/j.1742-4658.2006.05349.x. |
FEBS J |
Identification of a cowpea gamma-thionin with bactericidal activity
Abstract
- Antimicrobial peptides are an abundant group of proteinaceous compounds widely produced in the plant kingdom. Among them, the gamma-thionin family, also known as plant defensins, represents one typical family and comprises low molecular mass cysteine-rich proteins, usually cationic and distributed in different plant tissues. Here, we report the purification and characterization of a novel gamma-thionin from cowpea seeds (Vigna unguiculata), named Cp-thionin II, with bactericidal activity against Gram-positive and Gram-negative bacteria. Once the primary structure was elucidated, molecular modelling experiments were used to investigate the multimerization and mechanism of action of plant gamma-thionins. Furthermore, Cp-thionin II was also localized in different tissues in cowpea seedlings during germination in contrasting conditions, to better understand the plant protection processes. The use of plant defensins in the construction of transgenic plants and also in the production of novel drugs with activity against human pathogens is discussed.
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| 16826554 |
1H and 13C assignments of cyclo[N-(Lys-Phe)-Orn-Val], a semicyclic imide tetrapeptide from Burkholderia cepacia |
10.1002/mrc.1864. |
Magn Reson Chem |
1H and 13C assignments of cyclo[N-(Lys-Phe)-Orn-Val], a semicyclic imide tetrapeptide from Burkholderia cepacia
Abstract
- The rhizobacteria Burkholderia cepacia biosynthesized the new tetrapeptide Cyclo[N-(Lys-Phe)-Orn-Val] (1), a 2,5-diketopiperazine, and the known siderophore azurechelin (salicylic acid). The structure of 1 was established by means of IR, UV, 1H and 13C NMR, double dimension experiments and MS.
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