Construction of Chemical Libraries of Volatile Compounds by Combinatorial Synthesis of Homologous Mixtures: Alk-4-en-1-ols,Alk-4-enals and Methyl Alk-4-enoates

A compound library of sixty six linear compounds, eleven representatives of six molecular families: (E)- and (Z)-isomers of alk-4-en-1-ols, alk-4-enals, and methyl alk-4-enoates, was prepared by combinatorial syntheses to allow the creation of a mass spectral database directly usable for their identification in GC/MS analyses. We demonstrate here that compound libraries can be prepared by combinatorial syntheses using long linear synthetic sequences, i. e., eight step in the case of 4-enals. The resulting mixtures of homologues are still perfectly exploitable to deliver the requested information such as clean mass spectra and good gas chromatographic retention indices.     

Changes in protein synthesis during stratification and dormancy release in embryos of sugar maple (Acer saccharum)

Protein synthesis in dormant embryos of sugar maple ( Acer saccharum ) was investigated in seeds stratified at 4°C or incubated at 15°C. Seeds stratified at 4°C germinated after 27 days; seeds incubated at 15°C failed to germinate. Stratification increased the embryo's capacity for protein synthesis by day 11 as measured by in vivo incorporation of [³⁵S]-methionine into purified protein. At 4°C protein synthesis in the embryonic axis rose in a linear fashion prior to germination, whereas in cotyledons it increased until day 20 and then declined. Analysis of radiolabelled proteins by two-dimensional gel electrophoresis revealed that the levels of specific proteins were altered by temperature, primarily in the cotyledons. Several proteins were expressed in the cotyledons at 15°C but were absent in unstratified embryos and in embryos stratified at 4°C. That is, the expression of these proteins was repressed during stratification and release from dormancy. Levels of other proteins in the cotyledons declined at 4°C during stratification. We suggest that one or more of these proteins may be associated with the inhibition of growth of the embryonic axis imposed by the cotyledons.     

First chemical synthesis of a scorpion alpha-toxin affecting sodium channels: the Aah I toxin of Androctonus australis hector.

Aah I is a 63-residue alpha-toxin isolated from the venom of the Buthidae scorpion Androctonus australis hector, which is considered to be the most dangerous species. We report here the first chemical synthesis of Aah I by the solid-phase method, using a Fmoc strategy. The synthetic toxin I (sAah I) was renatured in DMSO-Tris buffer, purified and subjected to thorough analysis and comparison with the natural toxin. The sAah I showed physico-chemical (CD spectrum, molecular mass, HPLC elution), biochemical (amino-acid composition, sequence), immunochemical and pharmacological properties similar to those of the natural toxin. The synthetic toxin was recognized by a conformation-dependent monoclonal anti-Aah I antibody, with an IC50 value close to that for the natural toxin. Following intracerebroventricular injection, the synthetic and the natural toxins were similarly lethal to mice. In voltage-clamp experiments, Na(v) 1.2 sodium channel inactivation was inhibited by the application of sAah I or of the natural toxin in a similar way. This work describes a simple protocol for the chemical synthesis of a scorpion alpha-toxin, making it possible to produce structural analogues in time.     

Penicillin acylase-catalyzed synthesis of β-lactam antibiotics in water-methanol mixtures: effect of cosolvent content and chemical nature of substrate on reaction rates and yields

The synthesis of four β-lactam antibiotics (penicillin G, pivaloyloxymethyl ester of penicillin G, ampicillin and pivampicillin) catalyzed byEscherichia coli penicillin acylase has been investigated in water-methanol mixtures. The enzyme reactions were either thermodynamically or kinetically controlled at the same conditions using phenylacetic acid andd--phenylglycine methyl ester as acyl donors and 6-aminopenicillanic acid and pivaloyloxymethyl 6-aminopenicillanic acid as acyl acceptors. It has been found that the influences of the cosolvent content on the reaction rates and synthetic yields are significantly different depending on the substrates used in the experiments. On the other hand, within certain ranges of the methanol content (up to ca. 40% (v/v) the residual activities of the enzymes in water-methanol mixtures were only slightly lower than those in aqueous media. To analyze the factors that determine the reaction rate in water-cosolvent mixtures, the effect of methanol on the apparent pK values of the substrates has been investigated, and a mathematical model has been developed on the basis of the assumption that the enzyme binds non-ionized substrates. Model simulation results indicate that the solvent effect on reaction rates is mainly attributed to the kinetic effects of changes in apparent pK values.     

Protein Synthesis in Euglena gracilis is Light-and Temperature-Dependent,Oscillating in a Circadian,Temperature-Compensated Manner

Abstract: Incorporation of radiolabelled amino acids into proteins of Euglena gracilis revealed that the amount of labelled protein depends on the conditions of illumination and temperature of cultivation. Protein synthesis was generally lower under dark conditions except at 37 °C. The largest amounts of labelled protein were measured at 21 °C and decreased at higher and lower temperatures. By separating the labelled proteins of the membraneous cell fraction from subcultures under a range of culture conditions, the synthesis of some specific proteins was found to be light- and/or temperature-dependent. On incubating cells taken at different times during a light/dark cycle and under constant conditions, a circadian rhythm of ³⁵S-methionine- as well as ³⁵S-cysteine-incorporation was detected. Thereby the cells incorporated ten-times less cysteine than methionine. Protein synthesis always peaked during the last quarter of the daily light phase, confirming the rhythmic rise in total protein. The length of the rhythm period, approximately 24 h, was nearly independent of the applied temperature in the range of 16 to 27 °C.     

Mechanism of aluminium-induced porphyrin synthesis in bacteria

In previous studies, aluminium was found to retard bacterial growth and enhance porphyrin formation in Arthrobacter aurescens RS-2. The aim of this study was to establish the mechanism of action of aluminium which leads to increased porphyrin production. Cultures of Arthrobacter aurescens RS-2 were incubated in the absence and presence of 0.74 mm aluminium. After 6 and 24 h of incubation, various parameters of the haem biosynthetic pathway were determined. After 6 h of incubation with aluminium, the activities of the enzymes aminolevulinate synthase (ALAS), aminolevulinate dehydratase (ALAD), porphobilinogen deaminase (PBGD) and uroporphyrinogen decarboxylase (UROD) were increased by 120, 170, 190 and 203%, respectively, while that of ferrochelatase (FC) was found to be unchanged. However, after 24 h of incubation, no change in the activities of ALAS and ALAD was noted, while an about 2-fold increase in PBGD and UROD activities were observed. FC activity was decreased by 63%. It was concluded that aluminium exerts its effect by inducing the enzymes PBGD and UROD rather than by a direct or indirect effect on ALAS. Its effect on the final step in the haem biosynthetic pathway is discussed.     

Studies on epimerization-free methods for the preparation of aminosuccinyl peptides

Summary We have found that guanidine acetate catalyses the transformation of a -benzyl-aspartyl peptide (Boc-Asp-(OBzl)-Leu-Trp-OMe) to an aminosuccinyl peptide (Boc-Asu-Leu-Trp-OMe). The reaction was accompanied by partial epimerization. However, not even a small amount of epimerization could be detected when the aminosuccinyl peptide was synthesised from Boc-Asp-Leu-Trp-OMe with the addition of DIC, HOPfp and guanidine acetate (as a catalyst). This reaction seems to be suitable for the epimerization-free solid phase synthesis of aminosuccinyl peptides, e.g. Asu⁶-Lamprey-III-GnRH (Glp-His-Trp-Ser-His-Asu-Trp-Lys-Pro-Gly-NH₂).     

The role of nickel in acetyl-CoA synthesis by the bifunctional enzyme CO dehydrogenase/acetyl-CoA synthase: enzymology and model chemistry

 CO dehydrogenase/acetyl-CoA synthase (CODH/ACS) is one of the four known nickel enzymes. It is a bifunctional protein that catalyzes the oxidation of CO to CO₂ at a nickel iron-sulfur cluster (Cluster C) and a remarkable condensation reaction between a methyl group (donated from a methylated corrinoid iron-sulfur protein), carbon monoxide, and coenzyme A to form acetyl-CoA at a separate nickel iron-sulfur cluster (Cluster A). This review focuses on the current understanding of the structure and function of Cluster A and on related model chemistry. It describes studies that uncovered the first example of a biological organometallic reaction sequence. The mechanism of acetyl-CoA synthesis includes enzymebound methylnickel, iron-carbonyl, and acylmetal intermediates. Discovery of the methylnickel species constituted the first example of an alkylnickel species in biology and unveiled a new biological role for nickel. Received: 10 April 1996 / Accepted: 4 July 1996     

反义RNA技术在植物基因工程领域中的应用

反义RNA技术在植物基因工程领域中的应用包括：a．番茄和其他水果的成熟控制;b．植物的抗病性;c．改变花卉的颜色;d．植物淀粉合成的控制;e．油料植物种子中脂肪酸合成的控制;f．杂交种子生产中雄性不育性的控制;g．其他.     

硒对培养人胚肝细胞Ⅲ型前胶原,羟脯氨酸合成的影响

原代培养人胚肝细胞经１．１５６×１０￣(－７）ｍｏｌ／Ｌ硒预处理４ｈ,加入２０ｍｍｏｌ／Ｌ四氟化碳作用２０ｈ,观察硒对其Ⅲ型前胶原（ＰＣⅢ）和羟脯氨酸（Ｈｙｐ）生成的影响。结果培养液中ＰＣⅢ水平、细胞内Ｈｙｐ含量及细胞内外丙二醛（ＭＤＡ）水平均降低,与未加硒对照组比较差别有显著性（Ｐ＜０．０１）。而硒谷腕甘肽过氧化物酶（Ｓｅ－ＧＳＨ－ＰＸ）活性则较对照组显著增高（Ｐ＜０．００１）,且ＰＣⅢ水平与Ｓｅ－ＧＳＨ－Ｐ＿Ｘ／ＭＤＡ比值呈负相关（ｒ＝－０．９１５６,Ｐ＜０．０１）。提示硒可提高Ｓｅ－ＧＳＨ－Ｐ＿Ｘ／ＭＤＡ比值,抑制脂质过氧化激发的肝细胞胶原合成。