Mutational analysis of the functional domains of yeast K1 killer toxin.

To determine the functional domains of K1 killer toxin, we analyzed the phenotypes of a set of mutations throughout regions encoding the alpha- and beta-toxin subunits that allow secretion of mutant toxins. A range of techniques have been used to examine the ability of mutant toxins to bind to beta-glucan cell wall receptor and to form lethal ion channels. Our results indicate that both the alpha and beta subunits are involved in beta-glucan receptor binding. Defects in ion channel formation and toxin immunity are confined to the hydrophobic alpha subunit of the toxin.     

A monoclonal antibody that inhibits opioid binding to rat brain membranes

To understand the structure-function relationship and to probe the molecular characteristics of the purified opioid receptor, monoclonal antibodies (mab) were raised against a purified opioid receptor protein. After intensive screening of almost 1500 hybridoma cell lines, only 7 clones were shown to have very high immunoreactivity against the purified receptor. Moreover, out of these 7 clones, only 2, 3B4F11 and 3A27G, were found to inhibit the ligand binding property of the mu-opioid receptor. The mab 3B4F11 was found to inhibit 3H-diprenorphine binding to the purified receptor in a dose dependent manner with a maximal inhibition of 100% achieved with 20 micrograms of the antibody. Likewise, Fab fragments prepared from the mabs 3B4F11 inhibited 3H-diprenorphine binding to P2 membranes in a dose-dependent manner. In addition, it was found that the binding of 3H-DAGO, 3H-DPDPE and 3H-EKC was inhibited with approximately equal potency, suggesting that the Fabs prepared from the mab 3B4F11 interact with all 3 receptor types.     

The interaction of quinone analogues with wild-type and ubiquinone-deficient yeast mitochondria

The interaction of the exogenous quinones, duroquinone (DQ) and the decyl analogue of ubiquinone (DB) with the mitochondrial respiratory chain was studied in both wild-type and a ubiquinone-deficient mutant of yeast. DQ can be reduced directly by NADH dehydrogenase, but cannot be reduced by succinate dehydrogenase in the absence of endogenous ubiquinone. The succinate-driven reduction of DQ can be stimulated by DB in a reaction inhibited 50% by antimycin and 70-80% by the combined use of antimycin and myxothiazol, suggesting that electron transfer occurs via the cytochrome b-c1 complex. Both DQ and DB can effectively mediate the reduction of cytochrome b by the primary dehydrogenases through center o, but their ability to mediate the reduction of cytochrome b through center i is negligible. Two reaction sites for ubiquinol seem to be present at center o: one is independent of endogenous Q6 with a high reaction rate and a high Km; the other is affected by endogenous Q6 and has a low reaction rate and a low Km. By contrast, only one ubiquinol reaction site was observed at center i, where DB appears to compete with endogenous Q6. DB can oxidize most of the pre-reduced cytochrome b, while DQ can oxidize only 50%. On the basis of these data, the possible binding patterns of DB on different Q-reaction sites and the requirement for ubiquinone in the continuous oxidation of DQH are discussed.     

单面针的生物碱研究

自芸香科(Rutaceae)花椒属植物单面针(Zanthoxylum nitidum var. fastuosum How ex Huang)的根皮中分得五种已知生物碱:乙氧基白屈菜红碱(ethoxychelerythrine)(Ⅰ);氯化光花椒碱(nitidine chloride)(Ⅱ);去甲基白屈菜红碱(des-N-methychelerythrine)(Ⅲ);α—别隐品碱(α-allocryptopine)(Ⅳ);鹅掌揪宁(liriodenine)(Ⅴ).     

长叶车前花叶病毒(HRV_(sh))外壳蛋白赖氨酸残基的修饰

我们曾报道长叶车前花叶病毒上海分离株(简称HRVsh)的外壳蛋白有二个赖氨酸残基,在PH8.5无变性剂存在的条件下,完整病毒颗粒表面的赖氨酸残基可与三硝基苯磺酸(TNPS)起反应,反应后的TNP-HRVsh病毒颗粒的感染力丧失达90％以上。 本文又进行了甲基乙亚胺甲酯(MEI)对HRVsh赖氨酸残基的修饰反应,修饰后的MEI-HRVsh病毒颗粒的感染力也同样丧失90％以上。 从三硝基苯磺酸修饰的病毒颗粒(TNP-HRVsh)中分离得到的RNA能与天然的HRVsh的外壳蛋白重建病毒颗粒,并具有感染力,说明修饰过程中核酸并不受影响。 进一步用同位素~(35)S,~(32)P双标记病毒,再以TNPS修饰标记的病毒,得到(~(35)S,~(32)P)-HRVsh及TNP-(~(35)S,~(32)P)-HRVsh。将两者分别接种于系统寄主青菜(Brassica chinensis)的一片叶片,一天后在非接种叶片上都可测得~(35)S,~(32)P的放射计数。其中,(~(35)S,~(32)P)-HRVsh的~(35)S/~(32)P比值降低了,而TNP-(~(35)S,~(32)P)-HRVsh的~(35)S/~(32)P比值保持不变。说明HRVsh外壳蛋白赖氨酸残基的修饰并不影响病毒颗粒进入寄主细胞,以及在寄主细胞间的转移。同位素双标记的结果表明,其感染力丧失的原因可能是由于上述修饰作用阻止了病毒在感染中所必须的脱壳过程。     

绿茶抗氧化剂成分抑制突变作用的初步研究

绿茶水溶性提取物及茶叶中抗氧化剂成份具有明显的抑制AFB_1及Bap诱导的鼠伤寒沙门氏菌回复突变作用。这种抗氧化剂成份还可以抑制AFB_1和Bap诱导的V79细胞基因正向突变,以及AFB_1诱导的V79细胞SCE和染色体畸变。本实验结果提示,绿茶中抗氧化剂成份可能对AFB_1及Bap的致癌性具有抑制作用。本文就茶叶抗氧化剂抑制突变的可能机制进行了初步讨论。     

The complete amino-acid sequence of the proteinase inhibitor B from the root of the arrowhead (Sagittaria sagittifolia L.)

After reduction and alkylation of the disulfide bonds of the proteinase inhibitor B from the root of the arrowhead (Sagittaria sagittifolia L.) followed by CNBr cleavage three peptide fragments with 68, 62 and 11 amino-acid residues could be separated on DEAE-Sepharose CL-6B. The peptides or the inhibitor itself were further specifically cleaved either by trypsin or by the mixture of (CH3)2SO/HCl/HBr at the arginyl- and the tryptophyl-peptide bond, respectively. The complete amino-acid sequences of the peptides were determined by manual solid phase DABITC/PITC double coupling micro-method and the primary structure of the arrowhead inhibitor B consisting of 141 amino-acid residues was then elucidated. Twenty pairs of amino-acid residues are repeated in the molecule of this inhibitor, three of these pairs even occur three times. The possible locations of the reactive sites are discussed. On the basis of sequence comparisons between this inhibitor and all other serine proteinase inhibitors the arrowhead inhibitor may belong to a new family.     

兔边缘系统隔区呼吸相关神经元

本实验在42只家兔给与 Urethane 半量麻醉,在边缘系统隔区用玻璃微电极方法记录了60个自发的呼吸相关神经元单位放电:吸气型30个单位;呼气型16个单位;跨时相型14个单位。断双侧迷走神经,静脉注入尼克刹米后,呼吸相关神经元单位放电与呼吸节律变化具有伴随性,呈正相关。窒息可以诱发隔区神经元呼吸节律放电。延髓第四脑室局部注入2%Sod。pentothal 后,随呼吸节律抑制、隔区呼吸相关神经元单位放电立即消失。上述结果提示:到达边缘系统隔区的呼吸信息在自主功能及情绪活动的协调方面,可能被认为是有意义的。