| 17608619 |
Human alpha2-macroglobulin is composed of multiple domains, as predicted by homology with complement component C3 |
10.1042/BJ20070764. |
Biochem J |
Human alpha2-macroglobulin is composed of multiple domains, as predicted by homology with complement component C3
Abstract
- Human alpha2M (alpha2-macroglobulin) and the complement components C3 and C4 are thiol ester-containing proteins that evolved from the same ancestral gene. The recent structure determination of human C3 has allowed a detailed prediction of the location of domains within human alpha2M to be made. We describe here the expression and characterization of three alpha(2)M domains predicted to be involved in the stabilization of the thiol ester in native alpha2M and in its activation upon bait region proteolysis. The three newly expressed domains are MG2 (macroglobulin domain 2), TED (thiol ester-containing domain) and CUB (complement protein subcomponents C1r/C1s, urchin embryonic growth factor and bone morphogenetic protein 1) domain. Together with the previously characterized RBD (receptor-binding domain), they represent approx. 42% of the alpha2M polypeptide. Their expression as folded domains strongly supports the predicted domain organization of alpha2M. An X-ray crystal structure of MG2 shows it to have a fibronectin type-3 fold analogous to MG1-MG8 of C3. TED is, as predicted, an alpha-helical domain. CUB is a spliced domain composed of two stretches of polypeptide that flank TED in the primary structure. In intact C3 TED interacts with RBD, where it is in direct contact with the thiol ester, and with MG2 and CUB on opposite, flanking sides. In contrast, these alpha2M domains, as isolated species, show negligible interaction with one another, suggesting that the native conformation of alpha2M, and the consequent thiol ester-stabilizing domain-domain interactions, result from additional restraints imposed by the physical linkage of these domains or by additional domains in the protein.
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| 17609268 |
Fusion of cyclophilin A to Fv1 enables cyclosporine-sensitive restriction of human and feline immunodeficiency viruses |
10.1128/JVI.00616-07. |
J Virol |
Fusion of cyclophilin A to Fv1 enables cyclosporine-sensitive restriction of human and feline immunodeficiency viruses
Abstract
- TRIM5alpha is a potent intracellular antiviral restriction factor governing species-specific retroviral replication. In the New World species owl monkey the coding region for the viral binding B30.2 domain of TRIM5alpha has been replaced by a cyclophilin A (CypA) pseudogene by retrotransposition. The resultant TRIM5-CypA fusion protein restricts human immunodeficiency virus type 1 (HIV-1), as well as feline immunodeficiency virus (FIV), by recruitment of the CypA domain to the incoming viral capsids. Infectivity is rescued by agents such as cyclosporine that disrupt CypA binding to its substrates. Mice encode an antiviral restriction factor called Fv1 (for Friend virus susceptibility gene 1), which is active against murine leukemia virus and related to endogenous gag sequences. Here we show that fusing CypA to Fv1 generates a restriction factor with the antiviral specificity of TRIMCyp but the antiviral properties of Fv1. Like TRIMCyp, Fv1-Cyp restricts HIV-1 and FIV and is sensitive to inhibition by cyclosporine. TRIM5alpha is known to have a short half-life and block infectivity before viral reverse transcription. We show that Fv1-Cyp has a long half-life and blocks after reverse transcription, suggesting that its longer half-life gives the restricted virus the opportunity to synthesize DNA, leading to a later block to infection. This notion is supported by the observation that infectivity of Fv1-Cyp restricted virus can be rescued by cyclosporine for several hours after infection, whereas virus restricted by TRIMCyp is terminally restricted after around 40 min. Intriguingly, the Fv1-Cyp-restricted HIV-1 generates closed circular viral DNA, suggesting that the restricted virus complex enters the nucleus.
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| 17613531 |
Structure of substrate-free human insulin-degrading enzyme (IDE) and biophysical analysis of ATP-induced conformational switch of IDE. |
10.1074/jbc.m701590200 |
J. Biol. Chem. |
Structure of substrate-free human insulin-degrading enzyme (IDE) and biophysical analysis of ATP-induced conformational switch of IDE.
Abstract
- Insulin-degrading enzyme (IDE) is a zinc metalloprotease that hydrolyzes amyloid-beta (Abeta) and insulin, which are peptides associated with Alzheimer disease (AD) and diabetes, respectively. Our previous structural analysis of substrate-bound human 113-kDa IDE reveals that the N- and C-terminal domains of IDE, IDE-N and IDE-C, make substantial contact to form an enclosed catalytic chamber to entrap its substrates. Furthermore, IDE undergoes a switch between the closed and open conformations for catalysis. Here we report a substrate-free IDE structure in its closed conformation, revealing the molecular details of the active conformation of the catalytic site of IDE and new insights as to how the closed conformation of IDE may be kept in its resting, inactive conformation. We also show that Abeta is degraded more efficiently by IDE carrying destabilizing mutations at the interface of IDE-N and IDE-C (D426C and K899C), resulting in an increase in Vmax with only minimal changes to Km. Because ATP is known to activate the ability of IDE to degrade short peptides, we investigated the interaction between ATP and activating mutations. We found that these mutations rendered IDE less sensitive to ATP activation, suggesting that ATP might facilitate the transition from the closed state to the open conformation. Consistent with this notion, we found that ATP induced an increase in hydrodynamic radius, a shift in electrophoretic mobility, and changes in secondary structure. Together, our
Results highlight the importance of the closed conformation for regulating the activity of IDE and provide new molecular details that will facilitate the development of activators and inhibitors of IDE.
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| 17617692 |
Lariatins, novel anti-mycobacterial peptides with a lasso structure, produced by Rhodococcus jostii K01-B0171 |
10.1038/ja.2007.48. |
J Antibiot (Tokyo) |
Lariatins, novel anti-mycobacterial peptides with a lasso structure, produced by Rhodococcus jostii K01-B0171
Abstract
- Two anti-mycobacterial peptides with a lasso structure, named lariatins A and B, were separated by HP-20 and ODS column chromatographies and purified by HPLC from the culture broth of Rhodococcus jostii K01-B0171, which was isolated from soil aggregates collected in Yunnan, China. Lariains A and B showed growth inhibition against Mycobacterium smegmatis with MIC values of 3.13 and 6.25 microg/ml in agar dilution method, respectively. Furthermore, lariatin A inhibited the growth of Mycobacterium tuberculosis with an MIC of 0.39 microg/ml in liquid microdilution method.
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| 17620056 |
A novel horse alpha-defensin: gene transcription, recombinant expression and characterization of the structure and function |
10.1042/BJ20070747. |
Biochem J |
A novel horse alpha-defensin: gene transcription, recombinant expression and characterization of the structure and function
Abstract
- Defensins are a predominant class of antimicrobial peptides, which act as endogenous antibiotics. Defensins are classified into three distinct sub-families: theta-, beta-, and alpha-defensins. Synthesis of alpha-defensin has been confirmed only in primates and glires to date and is presumably unique for a few tissues, including neutrophils and Paneth cells of the small intestine. Antimicrobial activities of these peptides were shown against a wide variety of microbes including bacteria, fungi, viruses and protozoan parasites. In the present study, we report the characterization of the equine alpha-defensin DEFA (defensin alpha) 1. Transcription analysis revealed that the transcript of the gene is present in the small intestine only. An alignment with known alpha-defensins from primates and glires displayed a homology with Paneth-cell-specific alpha-defensins. DEFA1 was recombinantly expressed in Escherichia coli and subsequently analysed structurally by CD and molecular modelling. To examine the antimicrobial properties, a radial diffusion assay was performed with 12 different micro-organisms and the LD90 (lethal dose killing > or =90% of target organism) and MBC (minimal bactericidal concentration) values were examined. DEFA1 showed an antimicrobial activity against different Gram-positive and Gram-negative bacteria and against the yeast Candida albicans. Using viable bacteria in combination with a membrane-impermeable fluorescent dye, as well as depolarization of liposomes as a minimalistic system, it became evident that membrane permeabilization is at least an essential part of the peptide's mode of action.
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| 17623531 |
Gonadotrophin products: empowering patients to choose the product that meets their needs |
10.1016/s1472-6483(10)60688-8. |
Reprod Biomed Online |
Gonadotrophin products: empowering patients to choose the product that meets their needs
Abstract
- Gonadotrophins are injected daily over several days in follicular stimulation protocols. To facilitate self-injection, various injection or reconstitution devices are available. It was investigated whether injection training enabled patients to choose their preferred device. Patients and their partners received nurse-led training about: powdered urofollitropin (Bravelle) with needle-free reconstitution (Q-Cap)and conventional needles and syringes for administration; follitropin beta (Follistim AQ) in a premixed, prefilled cartridge (Follistim AQ Cartridge) with a reusable injection device (Follistim Pen); or follitropin alfa (GONAL-f) in a disposable, premixed, prefilled injection device (GONAL-f RFF Pen). A total of 123 participants (81 women) attended the training and were asked to complete a questionnaire after training. More participants expressed a preference for a pen injection device than the needle-free reconstitution and conventional syringe (84.6% versus 5.7%; P < 0.0001). Of the 94 participants who preferred a particular device, more preferred the follitropin alfa prefilled pen (68.1%) than the follitropin beta cartridge and pen (24.5%; P < 0.0001) or urofollitropin with needle-free reconstitution device and conventional syringe (7.4%; P < 0.0001). It was concluded that nurse-led training classes empowered participants to choose a device that they considered most suitable to their needs.
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| 17624760 |
The insulin-like growth factor 1 receptor in cancer: old focus, new future |
10.1016/j.ejca.2007.05.021. |
Eur J Cancer |
The insulin-like growth factor 1 receptor in cancer: old focus, new future
Abstract
- The importance of insulin-like growth factor 1 receptor (IGF-1R) signalling in malignant behaviour of tumour cells is well established. Currently, development of drugs targeting the IGF-1R as anticancer treatment is emerging. Several IGF-1R targeting strategies are being investigated in phases I and II clinical trials. Interactions of IGF-1R with insulin receptor, however, might complicate efficiency and tolerability of such drugs. This review describes mechanisms, recent developments and potential limitations of IGF-1R antibodies and tyrosine kinase inhibitors.
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| 17635581 |
Novel alpha-conotoxins from Conus spurius and the alpha-conotoxin EI share high-affinity potentiation and low-affinity inhibition of nicotinic acetylcholine receptors |
10.1111/j.1742-4658.2007.05931.x. |
FEBS J |
Novel alpha-conotoxins from Conus spurius and the alpha-conotoxin EI share high-affinity potentiation and low-affinity inhibition of nicotinic acetylcholine receptors
Abstract
- alpha-Conotoxins from marine snails are known to be selective and potent competitive antagonists of nicotinic acetylcholine receptors. Here we describe the purification, structural features and activity of two novel toxins, SrIA and SrIB, isolated from Conus spurius collected in the Yucatan Channel, Mexico. As determined by direct amino acid and cDNA nucleotide sequencing, the toxins are peptides containing 18 amino acid residues with the typical 4/7-type framework but with completely novel sequences. Therefore, their actions (and that of a synthetic analog, [gamma15E]SrIB) were compared to those exerted by the alpha4/7-conotoxin EI from Conus ermineus, used as a control. Their target specificity was evaluated by the patch-clamp technique in mammalian cells expressing alpha(1)beta(1)gammadelta, alpha(4)beta(2) and alpha(3)beta(4) nicotinic acetylcholine receptors. At high concentrations (10 microm), the peptides SrIA, SrIB and [gamma15E]SrIB showed weak blocking effects only on alpha(4)beta(2) and alpha(1)beta(1)gammadelta subtypes, but EI also strongly blocked alpha(3)beta(4) receptors. In contrast to this blocking effect, the new peptides and EI showed a remarkable potentiation of alpha(1)beta(1)gammadelta and alpha(4)beta(2) nicotinic acetylcholine receptors if briefly (2-15 s) applied at concentrations several orders of magnitude lower (EC(50), 1.78 and 0.37 nm, respectively). These results suggest not only that the novel alpha-conotoxins and EI can operate as nicotinic acetylcholine receptor inhibitors, but also that they bind both alpha(1)beta(1)gammadelta and alpha(4)beta(2) nicotinic acetylcholine receptors with very high affinity and increase their intrinsic cholinergic response. Their unique properties make them excellent tools for studying the toxin-receptor interaction, as well as models with which to design highly specific therapeutic drugs.
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| 17635696 |
Platelet integrin alpha(IIb)beta(3): activation mechanisms |
10.1111/j.1538-7836.2007.02537.x. |
J Thromb Haemost |
Platelet integrin alpha(IIb)beta(3): activation mechanisms
Abstract
- Integrin alpha(IIb)beta(3) plays a critical role in platelet aggregation, a central response in hemostasis and thrombosis. This function of alpha(IIb)beta(3) depends upon a transition from a resting to an activated state such that it acquires the capacity to bind soluble ligands. Diverse platelet agonists alter the cytoplasmic domain of alpha(IIb)beta(3) and initiate a conformational change that traverses the transmembrane region and ultimately triggers rearrangements in the extracellular domain to permit ligand binding. The membrane-proximal regions of alpha(IIb) and beta(3) cytoplasmic tails, together with the transmembrane segments of the subunits, contact each other to form a complex which restrains the integrin in the resting state. It is unclasping of this complex that induces integrin activation. This clasping/unclasping process is influenced by multiple cytoplasmic tail binding partners. Among them, talin appears to be a critical trigger of alpha(IIb)beta(3) activation, but other binding partners, which function as activators or suppressors, are likely to act as co-regulators of integrin activation.
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| 17643303 |
Synthesis and cytotoxicity of desmethoxycallipeltin B: lack of a qui methide for the cytotoxicity of callipeltin B |
10.1016/j.bmcl.2007.07.003. |
Bioorg Med Chem Lett |
Synthesis and cytotoxicity of desmethoxycallipeltin B: lack of a qui methide for the cytotoxicity of callipeltin B
Abstract
- The desmethoxy analogue of the cytotoxic, cyclic depsipeptide callipeltin B was synthesized to evaluate the role of its beta-MeOTyr residue. The IC(50) of desmethoxycallipeltin B, in which the beta-MeOTyr residue was replaced by d-Tyr, against HeLa cells was found to be 128+/-10 microM in an MTT assay, compared to 98+/-5 microM for the natural product itself. The roughly comparable cytotoxicities suggest that the cytotoxicity of callipeltin B does not arise through the formation of a qui methide intermediate.
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| 17653846 |
Efficacy of a combined protocol of urinary and recombinant follicle-stimulating hormone used for ovarian stimulation of patients undergoing ICSI cycle |
10.1007/s10815-007-9159-0. |
J Assist Reprod Genet |
Efficacy of a combined protocol of urinary and recombinant follicle-stimulating hormone used for ovarian stimulation of patients undergoing ICSI cycle
Abstract
- To evaluate the efficacy of using both urinary and recombinant FSH in a combined protocol for ovarian stimulation in an IVF treatment program.
A total of 119 infertile couples undergoing ICSI treatment were randomized prospectively in this study. After a standard down-regulation with GnRH analogue, the patients were randomized in 2 groups 58 received combined urinary and recombinant FSH, starting with uFSH and then rFSH, and 61 controls received only recombinant FSH.
Pregnancy and implantation rates were significantly higher in the combined uFSH/rFSH group than the control (rFSH) group (43.9% vs 22.1% and 27.5% vs 13.2% respectively). Metaphase II oocyte and grade 1 embryos were significantly higher in favour of combined uFSH/rFSH group than the recombinant FSH group.
This study shows that using a combination of both urinary and recombinant FSH for ovarian stimulation improves oocyte maturity and embryo cleavage, and increases pregnancy and implantation rates.
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| 17658600 |
Arasin 1, a proline-arginine-rich antimicrobial peptide isolated from the spider crab, Hyas araneus |
10.1016/j.dci.2007.06.002. |
Dev Comp Immunol |
Arasin 1, a proline-arginine-rich antimicrobial peptide isolated from the spider crab, Hyas araneus
Abstract
- Antimicrobial peptides (AMPs) are considered to play an important role as host-defense molecules in both vertebrates and invertebrates. This work was undertaken to characterize AMPs from hemocyte extracts of the small spider crab, Hyas araneus. A novel proline-arginine-rich AMP of 37 amino acids was isolated and characterized. The peptide, named arasin 1, has a chimeric structure with an N-terminal domain rich in proline and arginine and a C-terminal domain containing two disulfide linkages. The peptide precursor of 64 amino acids, deduced from a cDNA library, contained a hydrophobic pre-region of 25 amino acids, directly followed by the mature peptide. C-terminally, this precursor had two additional amino acids, which seem to be cleaved off post-translationally. Synthetic arasin 1 showed antibacterial activity. A putative isoform of arasin 1, named arasin 2, was found at the genetic level, and both transcripts were shown by real-time RT-PCR to be expressed mainly in hemocytes.
|
| 17660751 |
AChBP-targeted alpha-conotoxin correlates distinct binding orientations with nAChR subtype selectivity |
10.1038/sj.emboj.7601785. |
EMBO J |
AChBP-targeted alpha-conotoxin correlates distinct binding orientations with nAChR subtype selectivity
Abstract
- Neuronal nAChRs are a diverse family of pentameric ion channels with wide distribution throughout cells of the nervous and immune systems. However, the role of specific subtypes in normal and pathological states remains poorly understood due to the lack of selective probes. Here, we used a binding assay based on acetylcholine-binding protein (AChBP), a homolog of the nicotinic acetylcholine ligand-binding domain, to discover a novel alpha-conotoxin (alpha-TxIA) in the venom of Conus textile. Alpha-TxIA bound with high affinity to AChBPs from different species and selectively targeted the alpha(3)beta(2) nAChR subtype. A co-crystal structure of Ac-AChBP with the enhanced potency analog TxIA(A10L), revealed a 20 degrees backbone tilt compared to other AChBP-conotoxin complexes. This reorientation was coordinated by a key salt bridge formed between Arg5 (TxIA) and Asp195 (Ac-AChBP). Mutagenesis studies, biochemical assays and electrophysiological recordings directly correlated the interactions observed in the co-crystal structure to binding affinity at AChBP and different nAChR subtypes. Together, these results establish a new pharmacophore for the design of novel subtype-selective ligands with therapeutic potential in nAChR-related diseases.
|
| 17671655 |
Impaired basolateral sorting of pro-EGF causes isolated recessive renal hypomagnesemia |
10.1172/JCI31680. |
J Clin Invest |
Impaired basolateral sorting of pro-EGF causes isolated recessive renal hypomagnesemia
Abstract
- Primary hypomagnesemia constitutes a rare heterogeneous group of disorders characterized by renal or intestinal magnesium (Mg(2+)) wasting resulting in generally shared symptoms of Mg(2+) depletion, such as tetany and generalized convulsions, and often including associated disturbances in calcium excretion. However, most of the genes involved in the physiology of Mg(2+) handling are unknown. Through the discovery of a mutation in the EGF gene in isolated autosomal recessive renal hypomagnesemia, we have, for what we believe is the first time, identified a magnesiotropic hormone crucial for total body Mg(2+) balance. The mutation leads to impaired basolateral sorting of pro-EGF. As a consequence, the renal EGFR is inadequately stimulated, resulting in insufficient activation of the epithelial Mg(2+) channel TRPM6 (transient receptor potential cation channel, subfamily M, member 6) and thereby Mg(2+) loss. Furthermore, we show that colorectal cancer patients treated with cetuximab, an antagonist of the EGFR, develop hypomagnesemia, emphasizing the significance of EGF in maintaining Mg(2+) balance.
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| 17676801 |
Crotogossamide, a cyclic nonapeptide from the latex of Croton gossypifolius |
10.1021/np078007i. |
J Nat Prod |
Crotogossamide, a cyclic nonapeptide from the latex of Croton gossypifolius
Abstract
- A new cyclic nonapeptide, crotogossamide (1), was isolated from the latex of Croton gossypifolius. Its structure was elucidated by use of 1D and 2D NMR and MS and by hydrolysis followed by GC-MS analysis as cyclo(-Gly(1)-Ala(2)-Ser(3)-Gly(4)-Leu(5)-Asn(6)-Gly(7)-Ile(8)-Phe(9)-). This is the first report of a cyclic peptide from a Croton species. The known flavonoids kaempferol 3-O-rhamnopyranoside, quercitrin, and myricitrin were also isolated from the plant latex.
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