人工转录因子研究进展

转录因子是真核表达调控中非常重要的一类反式作用因子,通常由DNA结合结构域与效应结构域两部分组成,研究发现这两个结构域可以各自独立发生作用。基于转录因子的这种结构特点,可以人为地选择针对特定序列的DNA结合结构域与具有特定作用的效应结构域构建人工转录因子。目前人工转录因子的DNA结合结构域多为C₂H₂ 型锌指结构,每一个锌指单元由大约30个氨基酸组成,识别DNA双螺旋大沟中相连的3bp序列,并可通过氢键作用与相应的碱基结合;多个锌指可以串联成簇,从而识别并结合较长的DNA序列区域。常见的人工转录因子的效应结构域有激活结构域以及抑制结构域,不同的效应结构域赋予人工转录因子不同的功能。目前人工转录因子已经在基础研究、药物设计以及基因治疗等领域得到了广泛的应用。     

Maternal and neonatal scalp hair concentrations of zinc, copper, cadmium, and lead

Postpartum scalp hair samples from 82 term-pregnancy mother/neonate pairs were analyzed for their concentration of zinc (Zn), copper (Cu), cadmium (Cd), and lead (Pb), using inductively coupled plasma-mass spectrometry. Maternal and neonatal Zn concentrations had geometric means (and 99% confidence intervals) of 122.5 μg/g (117.9–131.5 μg/g) and 146.9 μg (141.5–156.7 μg/g) respectively. Corresponding Cu values were 18.4 μg/g (17.6–23.8 μg/g) and 6.7 μg/g (6.3–7.6 μg/g). Those of Cd were 0.49 μg/g (0.47–0.69 μg/g) in the mothers and 0.57 μg/g (0.55–0.86 μg/g) in the neonates. For Pb, they were 7.95 μg/g (7.60–9.32 μg/g) and 4.56 μg/g (4.39–5.56 μg/g). Cigaret smoking, despite its relatively low prevalence (19.5%), was associated with lower Zn and higher Cd and Pb concentrations and in lower Zn/Cd and Zn/Pb molar concentration ratios. Smoking also altered interelemental relationships, particularly those of Zn with Cd and Pb and those between Cd and Pb. Smoking frequency appeared to show negative dose-response effects on maternal and neonatal Zn concentrations, Zn/Pb molar concentration ratios, and birth weight. Mothers with a history of oral contraceptive (OC) usage had significantly higher Cu concentrations and lower Zn/Cu molar concentration ratios than nonusers, with the highest Cu concentrations and lowest Zn/Cu values being associated with third-generation OCs. No similar effects were elicited in the respective neonatal Cu concentrations. Neither alcohol consumption nor prenatal supplementation with iron and/or folic acid had discernible effects on the maternal or neonatal elemental concentrations. The data from this study suggest that in a given population of term-pregnancy mothers and neonates, significant interindividual variations in hair trace element concentrations can occur, irrespective of commonality of general environment, and that lifestyle factors, including cigaret smoking and OC usage history, can be significant contributory factors to such variations. The data are discussed in relation to the effects of smoking-associated exposure to Cd and Pb exposure on Zn availability for placental transfer, as well as on the quantitative maternal Zn supply levels to the fetus resulting from the known tendency of smokers to have lower dietary intakes of Zn. The higher Cu concentrations in OC users are discussed in relation to altered Cu metabolism, characterized by increased synthesis of the Cu-binding protein, ceruloplasmin, as an acute-phase antioxidant response to altered lipid profile and increased lipid oxidation.     

Mycorrhiza formation and nutrient concentration in leeks (Allium porrum) in relation to previous crop and cover crop management on high P soils

An improved integration of mycorrhizas may increase the sustainability in plant production. Two strategies for increasing the soil inoculum potential of mycorrhizal fungi were investigated in field experiments with leeks: Pre-cropping with mycorrhizal main crops and pre-establishment of mycorrhizal cover crops. Experiments on soils with moderate to high P content (26–50 mg kg^–1 bicarbonate-extractable P) showed that the previous crop influenced mycorrhiza formation, uptake of P, Zn, and Cu, and early growth of leek seedlings. A cover crop of black medic, established the previous autumn, increased the colonization of leek roots by mycorrhizal fungi. During early growth stages, this increase was 45–95% relative to no cover crop. However, cover cropping did not significantly increase nutrient concentration or growth. These variables were not influenced by the time of cover crop incorporation or tillage treatments. Differences in colonization, nutrient uptake and plant growth diminished during the growing period and at the final harvest date, the effects on plant production disappeared. High soil P level or high soil inoculum level was most likely responsible for the limited response of increased mycorrhiza formation on plant growth and nutrient concentrations.     

Modulation of zinc- and cobalt-binding affinities through changes in the stability of the zinc ribbon protein L36

Cysteine-rich Zn(II)-binding sites in proteins serve two distinct functions: to template or stabilize specific protein folds, and to facilitate chemical reactions such as alkyl transfers. We are interested how the protein environment controls metal site properties, specifically, how naturally occurring tetrahedral Zn(II) sites are affected by the surrounding protein. We have studied the Co(II)- and Zn(II)-binding of a series of derivatives of L36, a small zinc ribbon protein containing a (Cys)(3)His metal coordination site. UV-vis spectroscopy was used to monitor metal binding by peptides at pH 6.0. For all derivatives, the following trends were observed: (1) Zn(II) binds tighter than Co(II), with an average K (A) (Zn) /K (A) (Co) of 2.8(+/-2.0)x10(3); (2) mutation of the metal-binding ligand His32 to Cys decreases the affinity of L36 derivatives for both metals; (3) a Tyr24 to Trp mutation in the beta-sheet hydrophobic cluster increases K (A) (Zn) and K (A) (Co) ; (4) mutation in the beta-hairpin turn, His20 to Asn generating an Asn-Gly turn, also increases K (A) (Zn) and K (A) (Co) ; (5) the combination of His20 to Asn and Tyr24 to Trp mutations also increases K (A) (Zn) and K (A) (Co) , but the increments versus C(3)H are less than those of the single mutations. Furthermore, circular dichroism, size-exclusion chromatography, and 1D and 2D (1)H NMR experiments show that the mutations do not change the overall fold or association state of the proteins. L36, displaying Co(II)- and Zn(II)-binding sensitivity to various sequence mutations without undergoing a change in protein structure, can therefore serve as a useful model system for future structure/reactivity studies.     

镉锌复合污染对龙葵苗期生长和镉锌累积特性的影响

采用营养液培养法研究了苗期龙葵(Solanumnigrum L.)对镉锌复合污染的反应及其对镉锌的吸收和积累特性.结果表明:(1)高浓度镉(200μmol·L-1)、锌(500μmol·L-1)胁迫使苗期龙葵植株生长受到严重抑制,但低浓度镉锌复合处理的部分植株生物量甚至超过了对照.(2)添加锌对苗期龙葵地上部镉积累有显著的影响(P<0.05);当锌浓度为100μmol·L-1时,镉积累量达到峰值,其中的100Cd/100Znμmol·L-1处理组合的镉积累量为所有处理中的最大值(地上部积累量高达171.31μg·plant-1);当锌浓度增加到500μmol·L-1时,植株体内镉积累量则呈现下降趋势.(3)添加镉对苗期龙葵地上部锌积累也有显著的影响(P<0.05);当镉浓度达50μmol·L-1时,根、茎、叶中锌积累量均达到最高值;当镉浓度为200μmol·L-1时,各处理单株地上部锌积累量最低.(4)从植株整体的镉、锌积累量来看,苗期龙葵地上部镉积累量最高达根系的14.67倍,而其锌积累量最高达根系的13.59倍.研究发现,龙葵幼苗根系、茎和叶中镉、锌含量随营养液中镉、锌浓度的增加而显著增加(P<0.05);添加锌在一定程度上缓解了镉对苗期龙葵的毒害效应,低浓度锌还可以提高植株对镉的吸收和积累;添加镉在一定程度上增强了植株对锌的耐受性和吸收富集能力;植株地上部的镉、锌积累量明显高于根系.     

施锌对石灰性褐土上小白菜光合作用及保护酶活性的影响

通过田间试验的方法,采用30(低锌)、75(中锌)和120kg/hm2ZnSO4(高锌)3个施锌水平,研究施锌对山西省石灰性褐土上小白菜叶片叶绿素含量、净光合速率、硝酸还原酶(NR)活性、抗氧化酶活性及丙二醛(MDA)含量的影响.结果表明:随着施锌量的提高,小白菜叶片各叶绿素组分含量、总叶绿素含量、净光合速率和NR活性均呈先升后降的变化趋势,并均以中锌处理最大,且中、高锌处理显著高于对照(不施锌);小白菜叶片过氧化物酶(POD)和过氧化氢酶(CAT)活性也呈先升后降的趋势,而超氧化物歧化酶(SOD)活性持续增加,CAT、POD和SOD活性分别在低锌、中锌和高锌处理时达到最大值,所有施锌处理的3种保护酶活性均显著高于缺锌对照;各处理小白菜叶片MDA含量随着施锌量增大而逐渐降低,且所有施锌处理均显著低于缺锌对照.研究发现,施锌能有效增强石灰性褐土上小白菜叶片光合能力和氮同化能力,显著提高其抗氧化酶系活性,并以75kg/hm2ZnSO4效果最佳.     

Structural basis of the zinc- and terbium-mediated inhibition of ferroxidase activity in Dps ferritin-like proteins

Streptococcus suis Dpr is an iron-binding protein involved in oxidative stress resistance. It belongs to the bacterial Dps protein family whose members form dodecameric assemblies. Previous studies have shown that zinc and terbium inhibit iron incorporation in Listeria innocua Dps protein. In order to gain structural insights into the inhibitory effect of zinc and terbium, the crystal structures of Streptococcus suis Dpr complexes with these ions were determined at 1.8 A and 2.1 A, respectively. Both ions were found to bind at the ferroxidase center and in the same location as iron. In addition, a novel zinc-binding site formed by His40 and His44 was identified. Both His residues were found to be present within all known Streptococcus suis Dpr variants and in Streptococcus pneumoniae, Streptococcus gordonii, and Streptococcus sanguinis Dpr proteins. Amino acid sequence alignment of Dpr with other Dps family members revealed that His44 is highly conserved, in contrast to His40. The inhibitory effect of zinc and terbium on iron oxidation in Dpr was studied in vitro, and it was found that both ions at concentrations >0.2 mM almost completely abolish iron binding. These results provide a structural basis for the inhibitory effect of zinc and terbium in the Dps family of proteins, and suggest a potential role of the Dps proteins in zinc detoxification mechanisms involving the second zinc-binding site.     

Antibacterial activity of pyrrolidine dithiocarbamate

Pyrrolidine dithiocarbamate (PDTC), an antioxidant with a metal-chelating activity, has been used widely to inhibit the expression of inflammatory genes in vitro and in vivo. This study investigated whether PDTC has an antimicrobial activity against various bacteria. The antibacterial activity of PDTC and other compounds was evaluated in vitro by the broth microdilution method against Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, Staphylococcus aureus, and Escherichia coli. Bacterial growth was inhibited by PDTC, where a wide range of sensitivity was demonstrated among the tested bacteria. The antibacterial activity of PDTC was reduced by the addition of copper chloride; in contrast, it was enhanced considerably by zinc chloride. Two different zinc chelators, Ca-saturated EDTA (Ca-EDTA) and N,N,N',N'-tetrakis (2-pyridylmethyl) ethylenediamine, blocked the antibacterial activity of PDTC, whereas Zn-EDTA failed to reduce the activity of PDTC. These results demonstrate for the first time that PDTC possesses an antibacterial activity, for which zinc is required, and suggest that PDTC, possessing a dual anti-inflammatory and antibacterial activity, may be considered for topical use for inflammatory diseases of bacterial origin.