Protease-catalyzed semisynthesis of human neuropeptide Y

Human neuropeptide Y was semisynthesized by enzymatic condensation of des-Tyr36-NH2 human neuropeptide Y and H-Tyr-NH2 using Pseudomonas aeruginosa elastase, a metalloenzyme possessing a hydrolytic specificity for the imino side of hydrophobic amino acids. The optimum pH for this enzymatic synthesis was judged to be around 7 in a high concentration of an organic solvent.     

Solution synthesis of human peptide YY (hPYY)

Human peptide YY (hPYY) was synthesized in a conventional manner by assembling six peptide fragments followed by deprotection with 1 M trimethylsilyl trifluoromethanesulfonate (TMSOTf)-thioanisole in trifluoroacetic acid (TFA). After purification by gel-filtration on Sephadex G-25, followed by reversed-phase high-performance liquid chromatography, a highly purified sample of synthetic hPYY was obtained. When administered in dogs, synthetic hPYY was as active as synthetic porcine PYY in terms of the effects on systemic arterial blood pressure, and splanchnic blood flow.     

Practical synthesis of a neuropeptide Y antagonist via stereoselective addition to a ketene

[Reaction: see text]. The synthesis of neuropeptide Y antagonist 1, currently under clinical investigation for the treatment of obesity, is described. The convergent synthesis from trans-spirolactone carboxylic acid intermediate 2a and aminopyrazole 3 is predicated on a stereoselective route to the former. The coupling reaction of ethyl 4-oxocyclohexanecarboxylate (10a) with lithiated isonicotinamide 11 was investigated in detail, but even optimized conditions only provided a 45:55 ratio of trans:cis isomers (12a:12b). While selective crystallization schemes were developed to isolate the thermodynamically less stable trans isomer 2a, improved stereocontrol was subsequentially achieved by the application of ketene chemistry. The ketene formation and quench was investigated under a variety of conditions aimed at maximizing the trans:cis ratio. Reacting a mixture of carboxylic acids 2a and 2b with POCl3 in THF, followed by concomitant addition of tert-butyl alcohol in the presence of TMEDA at 35 degrees C provided a 4:1 ratio of trans:cis tert-butyl esters (18a:18b) via in situ ketene formation. Ester hydrolysis, followed by selective crystallization of undesired 2b as the HCl salt, led to isolation of 2a in 47% overall yield. Aminopyrazole intermediate 3 was synthesized via the condensation reaction of 2-fluorophenylhydrazine hydrochloride (4a) with acrylonitrile derivative 5 in 65-70% yield. Coupling of advanced intermediates 2a and 3b via activation with thionyl chloride gave a 92% yield of 1.     

Solution and gas-phase synthesis of the heteroligand yttrium complex with dipivaloylmethane and bis(salicylidene)ethylenediamine Y(dpm)(salen)

The interaction of yttrium dipivaloylmethanate Y(dpm)₃ with bis(salicylidene)ethylenediamine (H₂salen) in solution and the gas phase leads to the formation of the heteroligand complex Y(dpm)(salen). The composition of the product has been confirmed by IR spectroscopy, ¹H NMR, and elemental analysis. The introduction of a Schiff base excess into the solution or gas phase does not result in the formation of the homoleptic complex Y₂(salen)₃.     

Molecular modeling and dynamics of neuropeptide Y

Summary A combination of molecular modeling and molecular dynamics (MD) is used to determine a theoretical structure for neuropeptide Y (NPY). Starting with the X-ray structure for avian pancreatic polypeptide (APP), the substituted amino acids were mutated, the side chains oriented to local potential energy minima, and the entire structure minimized and subjected to an MD simulation. Comparison of the resulting NPY structure with APP X-ray and MD results showed secondary structural elements to be maintained and RMS fluctuations to be similar, although differences in both were observed. The approach presented offers a means to study the structure-function relationships of NPY and other similar polypeptides when combined with pharmacological measurements.Abbreviations NPY Neuropeptide Y - APP Avian pancreatic polypeptide - ABNR Adopted-basis Newton Raphson - MD Molecular dynamics     

Hormonal control of the neuropeptide Y system

Neuropeptide Y (NPY) and the related receptors represent a widely diffused system that is involved in the regulation of multiple biological functions. NPY, a 36-aminoacid peptide expressed in several areas of the nervous system, is a pleiotropic factor participating to the control of some physiological processes, such as cognitive functions, eating behavior, circadian rhythms, neuroendocrine mechanisms, reproductive and cardiovascular functions. NPY acts through a series of G-protein-associated membrane receptors (NPY-Rs), characterized by different tissue distribution and affinity for the ligand. The expression and secretion of NPY and the expression of NPY-R isoforms are controlled by a very wide range of agents, acting in an endocrine and/or paracrine fashion. NPY and NPY-Rs appear to be strongly involved in the control of eating behavior; their expression is modulated by changes of food intake and energy balance and is disrupted in several animal models of obesity and diabetes. Moreover, the hypothalamic NPY system appears to integrate signals of energy balance in the modulation of the reproductive axis. Agents that stimulate their expression include activators of intracellular signalling pathways (protein kinase A and C), classical neurotransmitters, steroid and peptide hormones and growth factors, while other agents (leptin, insulin and retinoic acid) have been shown to be inhibitory. Interestingly, some agents, like retinoic acid, have been shown to modulate the expression of both NPY and NPY-Rs in the same direction, thus providing a fine mechanism for the tuning of the system. The regulation of NPY/NPY-R expression and function appears to be part of a complex system controlling multiple physiological functions, and its disruption might be relevant in the pathophysiology of disease states such as obesity.     

芳氧基钇配合物引发丙烯腈溶液聚合的Monte Carlo模拟

用Monte Carlo方法模拟了DMF溶剂体系中单组分芳氧基钇引发剂Y(OAr)₃引发丙烯腈溶液聚合的全程反应动力学.通过计算机模拟和实验验证,表明聚合反应过程中存在配位阴离子活性中心,且活性中心容易形成,但很快失活.同时,存在向单体转移和向大分子转移两种链转移反应.获得了一组聚合反应动力学参数:k_i=0.053 L/(mol.m in),k_p=1.63 L/(mol.m in),k_d=0.005 8 m in^-1,k_tM=0.052 L/(mol.m in),k_trP=0.075 L/(mol.m in),k_pp=0 L/(mol.m in).     

Structure-affinity relationships of C-terminal cyclic analogue of neuropeptide Y for the Y1-receptor

We previously reported that a cyclic octapeptide amide, c[D-Cys29, Cys-34]NPY Ac-29-36 (YM-42454) showed a high affinity for Y1-receptors in SK-N-MC cells (Ki=0.047,microM) but not for Y2-receptors in the porcine hippocampus membranes (Ki>10microM). To explore the critical residues of this unique cyclic peptide for Y1-binding activity, the structure-affinity relationships were investigated by means of amino acid replacement. The results indicated that the hydrophobic side-chains of Leu30 and Ile31, the guanidinium groups of Arg33 and Arg33, and the C-terminal amide are critical for the binding affinity of YM-42454 to the Y1-receptor. On the other hand, Thr32 in YM-42454 might not be critical for the Y1-binding affinity. 1H-NMR studies for YM-42454 and its derivatives have suggested that the critical residues are involved in the direct interaction with a Y1-receptor rather than in maintaining the bioactive conformation.     

The synthesis of organoantimony(III) difluorides containing Y,C,Y pincer type ligands using organotin(IV) fluorinating agents

The organoantimony(III) difluorides containing Y,C,Y-chelating, so called pincer, ligands ([2,6-(YCH₂)₂C₆H₃]SbF₂; Y = MeO, t-BuO and Me₂N) were prepared by the reaction of corresponding dichlorides ([2,6-(YCH₂)₂C₆H₃]SbCl₂; Y = MeO, t-BuO and Me₂N) with two equivalents of organotin(IV) fluorinating agents Me₃SnF or 2-(Me₂NCH₂)C₆H₄Sn(n-Bu₂)F, respectively. The structure of organonantimony fluorides was determined both in solution by ¹H, ¹³C and ¹⁹F NMR spectroscopy and in the solid state using X-ray diffraction.     

Aptamer-functionalized graphene-gold nanocomposites for label-free detection of dielectrophoretic-enriched neuropeptide Y

We present an electrokinetically enhanced aptamer sensing platform on a disposable plastic chip for label-free detection of neuropeptide Y (NPY), which is a key neurological biomarker. The sensor consists of aptamer-functionalized graphene-gold nanocomposites (Gr-AuNs) patterned inside a nanoslit that is embossed on cyclic olefin copolymer via nanoimprint lithography. Analyte molecules are dielectrophoretically focused through the nanoslit onto aptamer-immobilized Gr-AuNs for rapid and selective electrochemical detection of NPY at picomolar levels.     

Solution and solid-phase chemoselective synthesis of (1-6)-amino(methoxy) di- and trisaccharide analogues

Disaccharide and trisaccharide mimics containing the amino(methoxy) interglycosidic linkage were obtained by chemoselective condensation of unprotected aldoses in an aqueous environment both in solution and in solid phase.     

Template synthesis and characterization of host (nanopores of zeolite Y)/guest (Co(II)-tetraoxodithiatetraaza macrocyclic complexes) nanocomposite materials

A series of Co(II) tetraoxodithiatetraaza macrocyclic complexes ([18]aneN₄S₂, [20]aneN₄S₂, Bzo₂[18]aneN₄S₂ and Bzo₂[20]aneN₄S₂) have been encapsulated in the nanopores of zeolite Y by template condensation reaction. Co(II) complexes with tetraoxodithiatetraaza macrocyclic ligand were entrapped in the nanopores of zeolite Y by a two-steps process in the liquid phase: (i) ion-exchange of [bis(diamine)cobalt(II)] (diamine = 1,2-diaminoethane, 1,3-diaminopropane, 1,2-diaminobenzene, 1,3-diaminobenzene); [Co(N–N)₂]²⁺–NaY; in the nano-cavity of the zeolite, and (ii) in situ template condensation of the cobalt(II) precursor complex with thiodiglycolic acid. The mode of bonding and overall geometry of the complexes and new host/guest nanocomposite materials ([Co([18]aneN₄S₂)]²⁺–NaY, [Co([20]aneN₄S₂)]²⁺–NaY, [Co(Bzo₂[18]aneN₄S₂)]²⁺–NaY, [Co(Bzo₂[20]aneN₄S₂)²⁺–NaY) has been inferred through FT-IR, DRS and UV–Vis spectroscopic techniques, BET technique, molar conductance and magnetic moment data, XRD and elemental analysis, as well as nitrogen adsorption. The average number of encapsulated Co complexes per nano-cavity was determined to be 0.33 for the Co complexes–NaY. An octahedral geometry around the cobalt(II) ion is suggested for the complexes and new host/guest nanocomposite materials.     

3D flower-like Y(2)O(3):Eu(3+) nanostructures: template-free synthesis and its luminescence properties

In this work, a facile route using simple hydrothermal reaction and sequential calcinations to synthesize 3-dimensional flower-like Y(2)O(3):Eu(3+) nanoarchitectures without employing templates or matrix for self-assembly is presented. The flower-like nanostructures are composed of nanosheets with thickness of about 30 nm, which is verified by the field-emission electron microscopy (FESEM). Influencing factors such as the dosage of reactants, the solvent, and pH are systematically investigated. The time-dependent experiments indicate a self-assembly mechanism. This method is also applicable in the preparation of other lanthanide oxides. The PL spectra of the as-synthesized Y(2)O(3):Eu(3+) are systematically studied. Both the Eu(3+) concentration and the calcinations temperature have great effect on the luminescence intensity of (5)D(0)-(7)F(2) transition. The decay curve of the (5)D(0) transition shows that the lifetime of the as-obtained Y(2)O(3):Eu(3+) is about 2.4 ms.     

In vitro selection of aptamers with affinity for neuropeptide Y using capillary electrophoresis

Capillary electrophoresis-systematic evolution of ligands by exponential enrichment (CE-SELEX) was used to select aptamers for neuropeptide Y (NPY). This is the first example of a CE-SELEX selection for aptamers that bind a target molecule smaller than itself. One of the limitations of CE-SELEX is that the aptamer must exhibit a significant mobility shift when it binds the target to facilitate fraction collection. Before this study, it was not clear if smaller targets would be capable of inducing a large enough shift in mobility for CE-SELEX to be successful. NPY is a 36-amino acid peptide (MW = 4272 g/mol), much smaller than the 80-base ssDNA used in the selection ( approximately 25 kDa). NPY binding aptamers with 300-1000 nM dissociation constants were obtained after only four rounds of selection. The specificity of the aptamers was tested using human pancreatic polypeptide (hPP). hPP is a 36-amino acid peptide with approximately 50% homology with NPY. Aptamers with up to 42-fold selectivity for NPY over hPP were observed.     

Solution synthesis of gadolinium nanoparticles

Gadolinium nanoparticles have been produced at subambient temperature by alkalide reduction. The nanoparticles display maxima in the temperature dependence of their magnetization, cooled in the absence of an applied external field, at T(max) of 5.0 and 17.5 K for unheated samples and samples annealed at 1000 degrees C for 4 h, respectively. Field cooled behavior deviates at temperatures slightly above T(max), increasing at lower temperature. Curie-Weiss law fits of the high-temperature data yield magnetic moments in close agreement with those expected for noninteracting Gd(3+) ions, suggesting that the behavior seen is due to a magnetic transition rather than superparamagnetism. Magnetization is linearly dependent on field at temperatures higher than 7-8 times T(max) and shows remanence-free hysteresis at lower temperature, suggesting metamagnetism. Some annealed samples show evidence of additional ferromagnetic interactions below approximately 170 K. Magnetic entropy curves generated from magnetization data are consistent with that expected for a paramagnet.     

f-Element/crown ether complexes 2. The synthesis and crystal structure of Y(NO3)3(12-crown-4)

Y(NO₃)₃(12-crown-4) was prepared via reaction of the crown ether and Y(NO₃)₃·nH₂O in acetonitrile. Y(NO₃)₃(12-crown-4) crystallizes in the monoclinic space groupP2₁/c witha=12.084(5),b=8.524(4),c=15.150(6) Å, =91.62(3)⁰ andD _calc=1.92 g cm^–3 forZ=4. The structure was refined by least-squares to a final conventionalR value of 0.105 using 1249 independent observed reflections [I3(I)]. The title compound is isostructural with its Eu(III) analog. The yttrium ion is ten-coordinate, bonded to three bidentate nitrate groups and to the four oxygens of the crown ether. The coordination polyhedron is best described as a 4A, 6B-extended dodecahedron. The Y-O(nitrate) and Y-O(ether) separations average 2.44(5) and 2.46(4) Å, respectively.Supplementary Data relating to this article are deposited with the British Library as Supplementary Publication No. SUP 82040 (10 pages).For Part 1 see reference [6]