小菜蛾对杀螟丹抗性遗传的研究

利用室内选育的抗杀螟丹小菜蛾Plutella xylostella (L.)品系和敏感品系研究了该品系的抗性遗传形式,结果表明,小菜蛾对杀螟丹的抗性形式为常染色体多基因遗传,并呈不完全显性。该品系对６种常用杀虫剂的抗性谱测定结果表明,对杀虫双有较严重的正交互抗性;对敌敌畏、杀扑磷有低度交互抗性;对溴氰菊酯、灭多威和叶蝉散等药剂无交互抗性。还发现该品系对杀螟丹的抗性与乙酰胆碱酯酶和羧酸酯酶活性无关。     

杀虫双和杀螟丹选育对小菜蛾抗药性的形成及其抗性机制

用杀虫双和杀螟丹在实验室以点滴法处理小菜蛾Plutella xylostella L.四龄幼虫,以连续继代药剂淘汰选育其抗药性。至35代,药剂汰选的小菜蛾对杀虫双和杀螟丹的抗药性较选育前正常品系分别提高了51倍和25倍。其抗药性的形成发展均呈S形,可认为已成为抗性品系。以有机磷、氨基甲酸酯、拟除虫菊酯及有机氮等11种杀虫剂测试抗杀虫双小菜蛾品系和抗杀螟丹小菜蛾品系对常用药剂的敏感度结果表明：对杀虫双、杀螟丹和杀虫环之间有较严重的正交互抗性;对敌敌艮、马拉硫磷和杀螟松有轻微交互抗性产生;对溴氰菊酯、氯氰菊酯、氯菊酯和灭多威、久效威等药剂更加敏感,呈负交互抗性。用聚丙烯酰胺凝胶电泳(PAGE)法测定表明,抗药性产生与特异性酯酶的形成有一定关系。用比色法和酸度法测定,抗性品系的乙酰胆碱酯酶(AchE)活性降低,羧酸酯酶(CarE)活性无差异。加增效剂Pb和SV₁：于四龄幼虫表皮,对抗杀虫双小菜蛾晶系分别有6.28及1.45倍的增效作用;对抗杀螟丹小菜蛾品系分别有4.85及1.39倍的增效作用,可见多功能氧化酶(MFO)为小菜蛾抗杀虫双和抗杀螟丹的重要因子。     

小菜蛾对杀虫双、杀螟丹抗性品系选育及其抗性消长规律的研究(英语)

用改进的蛭石萝卜苗法饲养小菜蛾,以点滴法处理对杀虫双、杀螟丹两种药剂淘汰选育的小菜蛾抗性品系和不以药剂处理的敏感品系的各项生物学特性与同期田间种群比较,各品系间差异不显著,均没有生长发育和繁殖上的不利情况.5年来药剂淘汰选育,杀虫双品系至F_(85)代其抗性增长178倍;杀螟丹品系F_(80)代其抗性增长87倍,均已形成高抗品系.两个抗性品系经停止药剂淘汰选育后5代,抗性倍数分别从167倍降至57倍和74倍降至16倍,说明这两个抗性品系的抗性是不稳定的,在没有药剂选择压力的情况下,开始几代抗性下降较快,当下降到一定水平后抗性便比较稳定,似乎很难恢复原有的敏感性.     

抗杀虫双小菜蛾和抗杀螟丹小菜娥酯酶同工酶的研究

用聚丙烯酰胺凝胶电泳法对抗杀虫双、抗杀螟丹的两个小菜蛾品系的酯酶同工酶研究结果表明：小菜蛾对杀虫双和杀螟丹的抗性与特异性酯酶同工酶的形成及部分非特异性酯酶同工酶活性的变化有关;杀虫双和杀螟丹在选育小菜蛾抗性的过程中对其酯酶同工酶的影响有很大的相似性。     

抗杀虫双小菜蛾和抗杀螟丹小菜蛾酯酶同工酶的研究

用聚丙烯酰胺凝胶电泳法对抗杀虫双、抗杀螟丹的两个小菜蛾品系的酯酶同工酶研究结果表明：小菜蛾对杀虫双和杀螟丹的抗性与特异酯酶同工酶的形成及部分非特异性酯酶同工酶活性的变化有关;杀虫和杀螟丹在选育小菜蛾抗性的过程中过程中对其酯酶同工酶的影响有很大的相似性。     

不同品系小菜蛾成虫脑突触体 蛋白质磷酸化的研究

对小菜蛾Plutella xylostella(L.)敏感品系、抗溴氰菊酯品系、抗杀虫双品系和抗杀螟丹品系的成虫脑突触体蛋白质磷酸化进行了研究比较。结果表明：蛋白质磷酸化在各个品系中的表现是不一样的。cAMP和钙加钙调蛋白对不同品系小菜蛾脑蛋白质磷酸化都有不同程度的刺激作用;3种杀虫剂均对各品系小菜蛾的磷酸化反应有影响,杀虫双、杀螟丹表现为抑制,溴氰菊酯表现为加强。这种影响在敏感品系中表现得比抗性品系中要强烈。     

小菜蛾对杀虫双和杀螟丹抗性的现实遗传力

利用室内选育119代的小菜蛾Plutella xylostella抗性品系,分析了小菜蛾对杀虫双和杀螟丹抗药性的现实遗传力,并对遗传力和杀虫剂的杀死率对抗性发展速率的影响进行了预测,结果表明,室内继代药剂选择119代后,小菜蛾对杀虫双和杀螟丹抗性的遗传力很低（h^2=0.03)。因此,小菜蛾对这两种药剂产生更高水平抗性的潜力不大,在选择的前半段（F30）,杀虫双和杀螟丹的抗性遗传力（分别为0.14t 0.11)高于选择后半段（F80)的遗传力（均为0.05),选择前、后半段的选择差数没有明显差别。即使在室内选择条件下,假如斜率=2.0(δp=0.5),h^2=0.05,群体中70%个体致死时要获得10倍的抗性,约需34代,田间条件下,由于等位基因频率的变化、环境变异等因素的影响,抗性增长10倍则需要更长的时间,因此田间条件下小菜蛾在短期内不易对杀虫双和杀螟丹产生高水平抗性。     

菜蛾对杀虫双和杀螟丹抗药性遗传的DNA随机扩增多态性研究

利用室内选育119代的抗杀虫双和抗杀螟丹小菜蛾Plutellaxylostella品系和敏感品系,通过反复回交建立近等基因系.用Operon公司合成的80个引物对抗杀虫双近等基因系和抗杀螟丹近等基因系小菜蛾基因组DNA进行PCR扩增;在抗杀虫双近等基因系中有3个引物分别产生了1条特异扩增带,2个引物分别产生了2条特异扩增带,1个引物产生3条特异扩增带;在抗杀螟丹近等基因系中,有4个引物分别产生了1条特异扩增带,1个引物产生了2条特异扩增带.在培育近等基因系的多次回交过程中,与抗性有关的遗传因子被逐步置换到敏感品系的基因组中,因此可以认为这些特异带与小菜蛾对杀虫双和杀螟丹的敏感性有关.     

小菜蛾对杀虫双和杀螟丹抗药性遗传的DNA随机扩增多态性研究

利用室内选育119代的抗杀虫和抗杀螟丹小菜蛾Plutella xylostella品系和敏感品系,通过反复回交建立近等基因系。用Operon公司全盛的80个引物对抗杀虫比近等基因系和抗杀螟丹近等基因系小菜蛾基因组DNA进行PCR扩增;在抗杀虫双近等基因系中3个引物分别产生了1条特异扩增带,2个引物分别产生了2条特异扩增带,1个引物产生3条特异拉增带;在抗杀螟丹近等基因系中,有4个引物分别产生了1条特异扩增带,1个引物产生了2条特异扩增带,在培育近等基因系的多次回交过程中,与抗性有关的遗传因子被逐步换到敏感品系的基因组中,因此可以认为这些特异带与小菜蛾对杀虫双和杀螟丹的敏感性有关。     

广东小菜蛾对苏芸金杆菌的抗性研究

广东省深圳,东莞、惠阳及博罗等供香港(以下简称供港)菜区小菜蛾对有机化学农药的抗性与广州内销菜区相近或稍高,对Bt杀虫剂的抗性则是供港菜区明显高于广州内销菜区。几种酶抑制剂TPP、SVl及Pb对Bt制剂无明显增效作用,可见小菜蛾对Bt制剂的抗性与酯酶和多功能氧化酶(MFO)的关系不大。用Bt制剂Dipel(大宝)连代选育小菜蛾敏感品系,选育18代,小菜蛾的抗性较选育前提高35倍。该抗性品系小菜蛾对个别菌株Bt及巴丹、杀虫双、速灭杀丁、万灵、敌敌畏等无交互抗性,而对昆虫生长调节抑制剂有轻微交互抗性。相反,用巴丹和杀虫双选育出的小菜蛾抗性品系对npel仍表现敏感。抗性品系小菜蛾在无触毒条件下饲养,抗性会自然减退,但不同类杀虫剂的抗性减退速率不尽相同。     

Seasonal variation in leaf glucosinolates and insect resistance in two types of Barbarea vulgaris ssp. arcuata

Leaves from natural populations of Barbarea vulgaris ssp. arcuata (Brassicaceae) in Denmark were examined for glucosinolate content and resistance to the crucifer specialist flea beetle Phyllotreta nemorum. Two types of the plant (P- and G-type) could be recognized. Leaves of the G-type contained the glucosinolates (only side chains mentioned): (S)-2-hydroxy-2-phenylethyl- (2S), indol-3-ylmethyl- (4) and in trace amount (R)-2-hydroxy-2-phenylethyl- (2R), 2-phenylethyl- (1) and 4-methoxyindol-3-ylmethyl- (5). Leaves of the P-type were dominated by 2R and 4, and had only trace amounts of 1, 2S, and 5 but contained in addition the previously unknown (R)-2-hydroxy-2-(4-hydroxyphenyl)ethyl- (3R). The epimer, (S)-2-hydroxy-2-(4-hydroxyphenyl)ethyl- (3S) was found in populations believed to be hybrids, and in B. orthoceras. 2S, 2R, desulfo 2S,-2R, -3S and -3R were isolated and identified by NMR and MS. Acylated glucosinolates or allylglucosinolate were not detected in leaves. The glucosinolate content in August was variable, 3-46 micromol/g dry wt, but was low in most populations, 3-15 micromol/g dry wt. In general, the glucosinolate content increased during the autumn, to 35-75 micromol/g dry wt in November. The G-type was resistant to neonate larvae of Phyllotreta nemorum in August and September (survival in 3-day bioassay typically 0%), and gradually lost the resistance in October and November (survival in 3-day bioassay 40-90%), and there was no correlation between glucosinolate content and resistance. Neither did glucosinolates explain the difference in resistance between the P-type (always susceptible) and the G-type (resistant in the summer season).     

Novel N6-substituted adenosine 5'-N-methyluronamides with high selectivity for human adenosine A3 receptors reduce ischemic myocardial injury

We recently reported the identification of a novel human adenosine A3 receptor-selective agonist, (2S,3S,4R,5R)-3-amino-5-[6-[5-chloro-2-(3-methylisoxazol-5-ylmethoxy)benzylamino]purin-9-yl]-4-hydroxytetrahydrofuran-2-carboxylic acid methylamide (CP-608,039), with 1,260-fold selectivity for the human A3 versus human A1 receptor (DeNinno et al., J Med Chem 46: 353-355, 2003). However, because the modest (20-fold) rabbit A3 receptor selectivity of CP-608,039 precludes demonstration of A3-mediated cardioprotection in rabbit models, we identified another member of this class, (2S,3S,4R,5R)-3-amino-5-[6-(2,5-dichlorobenzylamino)purin-9-yl]-4-hydroxytetrahydrofuran-2-carboxylic acid methylamide (CP-532,903), which both retained human A3 receptor selectivity (210-fold; human A3/human A1 Ki: 23/4,800 nM) and had improved rabbit A3 receptor selectivity (90-fold; rabbit A3/rabbit A1 Ki: 23/2,000 nM). Infarct size was measured in Langendorff hearts or in vivo after 30 min of regional ischemia and 120 min of reperfusion. Five-minute perfusion with CP-532,903 before ischemia-reperfusion elicited a concentration-dependent reduction in infarct size in isolated hearts (EC50: 0.97 nM; maximum reduction in infarct size: 77%, P < 0.05 vs. control). Furthermore, administration of CP-532,903 (150 nM) at reperfusion also significantly reduced infarct size by 64% (P < 0.05 vs. control), which was not different (P > or = 0.05) from the cardioprotection provided by the same concentration of drug given before ischemia. The selective rabbit A1 receptor antagonist BWA1433 did not affect CP-532,903-dependent cardioprotection. In vivo, CP-532,903 (1 mg/kg) reduced infarct size by 50% in the absence of significant hemodynamic effects (mean arterial pressure, heart rate, rate-pressure product). CP-532,903 and CP-608,039 represent a novel class of human A3 receptor-selective agonists that may prove suitable for investigation of the clinical cardioprotective efficacy of A3 receptor activation.     

Enantiomers of cis-constrained and flexible 2-substituted GABA analogues exert opposite effects at recombinant GABA(C) receptors

The effects of the enantiomers of a number of flexible and cis-constrained GABA analogues were tested on GABA(C) receptors expressed in Xenopus laevis oocytes using two-electrode voltage-clamp electrophysiology. (1S,2R)-cis-2-Aminomethylcyclopropane-1-carboxylic acid ((+)-CAMP), a potent and full agonist at the rho1 (EC(50) approximately 40 microM, I(max) approximately 100%) and rho 2 (EC(50) approximately 17 microM, I(max) approximately 100%) receptor subtypes, was found to be a potent partial agonist at rho3 (EC(50) approximately 28 microM, I(max) approximately 70%). (1R,2S)-cis-2-Aminomethylcyclopropane-1-carboxylic acid ((-)-CAMP), a weak antagonist at human rho1 (IC(50) approximately 890 microM) and rho2 (IC(50) approximately 400 microM) receptor subtypes, was also found to be a moderately potent antagonist at rat rho3 (IC(50) approximately 180 microM). Similarly, (1R,4S)-4-aminocyclopent-2-ene-1-carboxylic acid ((+)-ACPECA) was a full agonist at rho1 (EC(50) approximately 135 microM, I(max) approximately 100%) and rho2 (EC(50) approximately 60 microM, I(max) approximately 100%), but only a partial agonist at rho3 (EC(50) approximately 112 microM, I(max) approximately 37%), while (1S,4R)-4-aminocyclopent-2-ene-1-carboxylic acid ((-)-ACPECA) was a weak antagonist at all three receptor subtypes (IC(50)>300 microM). 4-Amino-(S)-2-methylbutanoic acid ((S)-2MeGABA) and 4-amino-(R)-2-methylbutanoic acid ((R)-2MeGABA) followed the same trend, with (S)-2MeGABA acting as a full agonist at the rho1 (EC(50) approximately 65 microM, I(max) approximately 100%), and rho2 (EC(50) approximately 20 microM, I(max) approximately 100%) receptor subtypes, and a partial agonist at rho3 (EC(50) approximately 25 microM, I(max) approximately 90%). (R)-2MeGABA, however, was a moderately potent antagonist at all three receptor subtypes (IC(50) approximately 16 microM at rho1, 125 microM at rho2 and 35 microM at rho3). On the basis of these expanded biological activity data and the solution-phase molecular structures obtained at the MP2/6-31+G* level of ab initio theory, a rationale is proposed for the genesis of this stereoselectivity effect.     

2-Amino-3-(3-hydroxy-1,2,5-thiadiazol-4-yl)propionic acid: resolution, absolute stereochemistry and enantiopharmacology at glutamate receptors

In order to identify new subtype-selective (S)-glutamate (Glu) receptor ligands we have synthesized (RS)-2-amino-3-(3-hydroxy-1,2,5-thiadiazol-4-yl)propionic acid [(RS)-TDPA]. Resolution of (RS)-TDPA by chiral chromatography was performed using a Crownpac CR(+) column affording (R)- and (S)-TDPA of high enantiomeric purity (enantiomeric excess=99.9%). An X-ray crystallographic analysis revealed that the early eluting enantiomer has R-configuration. Both enantiomers showed high affinity as well as high agonist activity at (RS)-2-amino-3-(3-hydroxy-5-methylisoxazol-4-yl)propionic acid (AMPA) receptors, determined using a [(3)H]AMPA binding assay and an electrophysiological model, respectively. The affinities and agonist activities obtained for (R)-TDPA (IC(50)=0.265 microM and EC(50)=6.6 microM, respectively) and (S)-TDPA (IC(50)=0.065 microM and EC(50)=20 microM, respectively) revealed a remarkably low AMPA receptor stereoselectivity, (S)-TDPA showing the highest affinity and (R)-TDPA the most potent agonist activity. In addition, (S)-TDPA was shown to interact with synaptosomal Glu uptake sites displacing [(3)H](R)-aspartic acid (IC(50 ) approximately 390 microM). An enantiospecific and subtype-selective agonist activity was observed for (S)-TDPA at group I metabotropic Glu (mGlu) receptors (EC(50)=13 microM at mGlu(5) and EC(50)=95 microM at mGlu(1)).     

Evidence for recent as well as long term activation of T cells migrating through endothelial cell monolayers in vitro.

As T cells actively extravasate from blood, they adhere to endothelium and then migrate out of the vessel with a locomotive activity. Although both adhesion and locomotion are properties associated with activated T cells, the two processes are not necessarily associated with identical activation states. Using human endothelial cells (EC) cultured to confluence on collagen gel, we examined the activation state of human peripheral blood T cells that adhere to and migrate through EC monolayers with three different methods: flow cytometric analysis of cell surface activation-related molecules, incorporation of tritiated nucleotide, and cell cycle analysis. The results were as follows. 1) Although expression of very late activation Ag integrins VLA-2 and VLA-3 by the initial blood T cell population (unseparated cells) and of adherent T cells was minimal, 40 to 45% of migrating cells were positive for VLA-2 and VLA-3. 2) The percentage of IL-2R+ cells in both unseparated and adherent cells was below 5% whereas the percentage of IL-2R+ cells among the migrating cells was 22 +/- 9% (range, 12 to 31%, n = 6). 3) Migrating cells expressed the highest CD26, whereas CD26 of adherent (nonmigrating) cells was divided into negative and high expression; in contrast, leukocyte adhesion molecule-1 (L-selectin) of both adherent and migrating cells was mostly low or negative. 4) [3H]Uridine incorporation of migrating and adherent cells was 2.1- to 2.5-fold and 1.4- to 1.7-fold higher, respectively, than that of unseparated cells, indicating that RNA synthesis of migrating cells as well as adherent cells was enhanced. 5) Cell cycle analysis showed that 23.5% of migrating cells appeared to enter the G1 phase but not S or G2 + M phases whereas 2.2% of unseparated cells and 8.0% of adherent cells that did not migrate had an RNA content consistent with entry into G1. These results suggest that cells migrating from normal human blood through unactivated EC have been activated recently as well as showing evidence of long term activation. The activation state of migrating cells is consistent with the hypothesis that previous in vivo activation is required for cells to migrate through EC in this system.     

Arg-274 and Leu-277 of the gamma-aminobutyric acid type A receptor alpha 2 subunit define agonist efficacy and potency

Alanine-scanning mutagenesis and the whole cell voltage clamp technique were used to investigate the function of the extracellular loop between the second and third transmembrane domains (TM2-TM3) of the gamma-aminobutyric acid type A receptor (GABA(A)-R). A conserved arginine residue in the TM2-TM3 loop of the GABA(A)-R alpha(2) subunit was mutated to alanine, and the mutant alpha(2)(R274A) was co-expressed with wild-type beta(1) and gamma(2S) subunits in human embryonic kidney (HEK) 293 cells. The GABA EC(50) was increased by about 27-fold in the mutant receptor relative to receptors containing a wild-type alpha(2) subunit. Similarly, the GABA EC(50) at alpha(2)(L277A)beta(1)gamma(2S) and alpha(2)(K279A)beta(1)gamma(2S) GABA(A)-R combinations was increased by 51- and 4-fold, respectively. The alpha(2)(R274A) or alpha(2)(L277A) mutations also reduced the maximal response of piperidine-4-sulfonic acid relative to GABA by converting piperidine-4-sulfonic acid into a weak partial agonist at the GABA(A)-R. Based on these results, we propose that alpha(2)(Arg-274) and alpha(2)(Leu-277) are crucial to the efficient transduction of agonist binding into channel gating at the GABA(A)-R.     

Mutation in the UL97 open reading frame of human cytomegalovirus strains resistant to ganciclovir.

The same point mutation in the human cytomegalovirus UL97 open reading frame was found in three independently isolated ganciclovir-resistant mutants of strain AD169. Point mutations in the DNA polymerase genes of these strains have been previously identified (N.S. Lurain, K.D. Thompson, E.W. Holmes, and G.S. Read, J. Virol. 66:7146-7152, 1992). All three strains are, therefore, double mutants. To determine the contribution of the UL97 mutation to the high ganciclovir resistance of these mutants, the mutation from the ganciclovir-resistant strain D6/3/1 was transferred to the wild-type strain AD169 to produce the recombinant R6HS. The ganciclovir resistance of R6HS is 4-fold lower than that of D6/3/1 but 10-fold higher than that of AD169. R6HS, like AD169, is sensitive to the nucleotide analogs (S)-1-[(3-hydroxy-2-phosphonylmethoxy) propyl]adenine and (S)-1-[(3-hydroxy-2-phosphonylmethoxy)propyl]cytosine. Ganciclovir phosphorylation in R6HS-infected cells was at the same reduced level as that found in cells infected with the parental mutant D6/3/1. The same G-to-T transversion at nucleotide 1380 in the UL97 coding sequence is present in both R6HS and D6/3/1. This mutation results in the substitution of isoleucine for methionine at amino acid residue 460. In an alignment of the R6HS UL97 amino acid sequence with the amino acid sequences of a wide range of protein kinase family members, methionine 460 lies within a highly conserved region which may function in nucleotide binding and phosphate transfer.     

Anti-HIV benzylisoquinoline alkaloids and flavonoids from the leaves of Nelumbo nucifera, and structure-activity correlations with related alkaloids

(+)-1(R)-Coclaurine (1) and (-)-1(S)-norcoclaurine (3), together with quercetin 3-O-beta-D-glucuronide (4), were isolated from the leaves of Nelumbo nucifera (Nymphaceae), and identified as anti-HIV principles. Compounds 1 and 3 demonstrated potent anti-HIV activity with EC50 values of 0.8 and <0.8 microg/mL, respectively, and therapeutic index (TI) values of >125 and >25, respectively. Compound 4 was less potent (EC50 2 microg/mL). In a structure-activity relationship study, other benzylisoquinoline, aporphine, and bisbenzylisoquinoline alkaloids, including liensinine (14), negferine (15), and isoliensinine (16), which were previously isolated from the leaves and embryo of Nelumbo nucifera, were evaluated for anti-HIV activity. Compounds 14-16 showed potent anti-HIV activities with EC50 values of <0.8 microg/mL and TI values of >9.9, >8.6, and >6.5, respectively. Nuciferine (12), an aporphine alkaloid, had an EC50 value of 0.8 microg/mL and TI of 36. In addition, synthetic coclaurine analogs were also evaluated. Compounds 1, 3, 12, and 14-16 can serve as new leads for further development of anti-AIDS agents.     

褐飞虱抗甲胺磷品系的交互抗性和抗性生化机制

用甲胺磷筛选的褐飞虱Nilaparvata lugens品系（R）,对甲胺磷的抗性达到43.74倍,对马拉硫磷、二嗪磷、异丙威、仲丁威及醚菊酯都表现出一定的交互抗性,而对氰戊菊酯和吡虫啉的交互抗性不显著。为了研究褐飞虱对甲胺磷抗性和对其它药剂交互抗性产生的机制,进行了活体增效试验和离体生化实验。用2 μg/头的增效剂预处理试虫的活体增效实验结果显示,在甲胺磷筛选品系（R）中, TPP(triphenyl phosphate, 磷酸三苯酯)对甲胺磷的增效倍数达到4.54,TPP对马拉硫磷、二嗪磷、仲丁威、异丙威都表现出一定的增效作用,增效比分别为2.76、2.07、2.17和1.64;PBO(piperonyl butoxide,胡椒基丁醚)对甲胺磷、马拉硫磷和醚菊酯有一定的增效作用;DEM(diethyl meteate, 顺丁烯二酸二乙酯)的增效作用不明显。研究离体情况下增效剂对三种解毒酶活性的影响发现,TPP对R品系酯酶活力抑制作用很强（抑制率69.04%）,PBO对多功能氧化酶（MFO）具有一定的抑制作用（抑制率29.30%）,而TPP和PBO在F品系和S品系中对酯酶和MFO的抑制作用都较小;DEM在三个品系中对谷胱甘肽-S-转移酶的抑制作用都很小。由此可见,酯酶在褐飞虱对甲胺磷的抗性中起最主要作用,在马拉硫磷、二嗪磷、异丙威和仲丁威的交互抗性中起很重要作用;MFO可能在甲胺磷抗性和醚菊酯、马拉硫磷的交互抗性中起一定作用。     

Purification and properties of two oxidoreductases catalyzing the enantioselective reduction of diacetyl and other diketones from baker's yeast

The NADPH-linked diacetyl reductase system from the cytosolic fraction of Saccharomyces cerevisiae has been resolved into two oxidoreductases catalyzing irreversibly the enantioselective reduction of diacetyl (2,3-butanedione) to (S)- and (R)-acetoin (3-hydroxy-2-butanone) [so-called (S)- and (R)-diacetyl reductases] (EC 1.1.1.5) which have been isolated to apparent electrophoretical purity. The clean-up procedures comprising streptomycin sulfate treatment, Sephadex G-25 filtration, DEAE-Sepharose CL-6B column chromatography, affinity chromatography on Matrex Gel Red A and Superose 6 prep grade filtration led to 120-fold and 368-fold purifications, respectively. The relative molecular mass of the (R)-diacetyl reductase, estimated by means of HPLC filtration on Zorbax GF 250 and sodium dodecyl sulfate/polyacrylamide gel electrophoresis, was 36,000. The (R)-enzyme was most active at pH 6.4 and accepted in addition to diacetyl C5-, C6-2,3-diketones, 1,2-cyclohexanedione, 2-oxo aldehydes and short-chain 2- and 3-oxo esters as substrates. The enzyme was characterized by high enantioselectivity and regiospecificity. The Km values for diacetyl and 2,3-pentanedione were determined as 2.0 mM. The Mr of the (S)-diacetyl reductase was determined as 75,000 by means of HPLC filtration of Zorbax GF 250. The enzyme decomposed into subunits of Mr 48,000 and 24,000 on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The optimum pH was 6.9. The purified (S)-enzyme reduced stereospecifically a broad spectrum of substrates, comprising 2,3-, 2,4- and 2,5-diketones, 2-oxo aldehydes, 1,2-cyclohexanedione and methyl ketones as well as 3-, 4- and 5-oxo esters. The 2,3- and 2,4-diketones are transformed to the corresponding (S)-2-hydroxy ketones; 2,5-hexanedione, however, was reduced to (S,S)-2,5-hexanediol. The Km values for diacetyl and 2,3-pentanedione were estimated as 2.3 and 1.5 mM, respectively. Further characterization of the (S)-diacetyl reductase revealed that it is identical with the so-called '(S)-enzyme', involved in the enantioselective reduction of 3-, 4- and 5-oxo esters in baker's yeast.