Mechanism of promoter repression by Lac repressor–DNA loops

The Escherichia coli lactose (lac) operon encodes the first genetic switch to be discovered, and lac remains a paradigm for studying negative and positive control of gene expression. Negative control is believed to involve competition of RNA polymerase and Lac repressor for overlapping binding sites. Contributions to the local Lac repressor concentration come from free repressor and repressor delivered to the operator from remote auxiliary operators by DNA looping. Long-standing questions persist concerning the actual role of DNA looping in the mechanism of promoter repression. Here, we use experiments in living bacteria to resolve four of these questions. We show that the distance dependence of repression enhancement is comparable for upstream and downstream auxiliary operators, confirming the hypothesis that repressor concentration increase is the principal mechanism of repression loops. We find that as few as four turns of DNA can be constrained in a stable loop by Lac repressor. We show that RNA polymerase is not trapped at repressed promoters. Finally, we show that constraining a promoter in a tight DNA loop is sufficient for repression even when promoter and operator do not overlap.     

半理性改造提升细菌漆酶Lac15的催化活性

[目的]漆酶可氧化各种底物,在多个工业领域有很好的潜在应用价值.Lac15是一种微生物漆酶,已表现出可观的应用潜能,可望通过蛋白质工程改造提升和拓展其应用.[方法]通过基于结构分析的半理性改造策略,选取推测与电子/质子转移或底物结合直接或间接相关的位点进行定点突变,并测定突变酶对各种底物的活性及酶学性质.[结果]部分突...     

重组海洋细菌漆酶Lac15脱色人工合成纺织染料的潜力

以重组海洋细菌漆酶Lac15(rLac15)为生物催化剂,对蒽醌类和偶氮类染料进行脱色,考察了rLac15对人工合成纺织染料的脱色潜能。通过研究催化的介体、酶量、pH、染料浓度以及温度对脱色效率的影响,进一步优化了rLac15对部分蒽醌类和偶氮类人工合成纺织染料的脱色条件。以丁香酸甲酯为介体,在pH 8.5、45℃条件下反应1 h,20 U/L rLac15对100μmol/L偶氮染料类Acid Red 6B(AR-6B),Reactive Blue 194(M-2GE),Reactive Brilliant Orange(K-7R)和Reactive Blue 171(KE-R)具有较好的脱色效果,脱色率分别达到95%、93%、76%和61%。随着染料浓度的增加,脱色率呈下降趋势,但当染料浓度达到200μmol/L时,M-2GE和AR-6B仍可保持80%以上脱色率。在常温下,rLac15对AR-6B、M-2GE、K-7R和KE-R显示较高脱色率,25℃反应24 h,分别达到96%、86%、66%和66%。rLac15是具有常温以及偏碱性环境脱色能力的细菌漆酶,具有潜在工业应用价值。     

High concentrations of viruses in the sediments of Lac Gilbert, Québec

Viruses were found to be very abundant in the top layer of the sediments of Lac Gilbert, Québec. Viruses were extracted from the sediments using pyrophosphate buffer, and viruses from the diluted extracts were pelleted onto grids and enumerated using transmission electron microscopy. Viral abundance in the sediments ranged from 6.5 × 10⁸ to 1.83 × 10¹⁰ ml^–1, which is 10- to 1,000-fold greater than the number observed in the water column. This increase corresponds well with the 100- to 1,000-fold increase in bacterial abundance in the sediments. Viral abundance differed significantly among the surface sediment samples taken at different bottom depths and among samples taken at different depths of the water column. Viral abundance also varied significantly between the oxic and anoxic zones of the water column and the sediments. The virus-to-bacteria ratio varied greatly among the different sediment sites but not among depths in the water column. Viral abundance in the water column was related to bacterial abundance and chlorophyll concentration, whereas viruses in the sediments were most abundant in sediments with high organic matter content. Elevated viral abundance and their erratic distribution in the sediments suggest that viruses might play an important role in sediment microbial dynamics. Correspondence to: Roxane Maranger     

The manganese cycle in Lac Léman,Switzerland: the role ofMetallogenium

The manganese pathways in Lac Léman have been investigated on the basis of chemical analyses undertaken on water, suspended solids, bottom sediments and interstitial water samples. The various modes of occurrence of Mn have been determined by means of visual examination using SEM and TEM (scanning and transmission electron microscopy), by microanalysis (EDAX) of various sediment fractions, by chemical analysis of the dissolved phase, and by chemical speciation and XRD of bottom sediments. Fluxes to and from the sediment have also been computed. The time-depth variations of Mn in the water column are characterized by (a) a very steep gradient of Mn sol. from the sediment interstitial water (15 mg l⁻¹) to the overlying water, 2 m above the bottom(500 μg l⁻¹), (b) an accumulation of Mn part. between 280 m and the interface at 310 m, consisting of mineralized colonies ofMetallogenium. The abundance ofMetallogenium colonies is inversely related to O₂ concentration; the optimal value for the bacterium Mn fixation is around 1 mg l⁻¹. Because of the quasi-anoxic state of the bottom sediments and overlying water, Mn diffuses from a ‘source layer’, 2–5 cm below the interface (a) upwards across the interface, before being taken up byMetallogenium, and (b) downwards to a ‘sink layer’, in which large amounts of Mn-carbonate precipitate. Particulate Mn sedimentation rates measured in traps show that downwards Mn flux due toMetallogenium settling approximately balances the upwards soluble flux due to diffusion. Quantitatively, the process of Mn cycling in Lac Léman is, therefore, limited to the lowermost part of the hypolimnion. Although Zn and Cd seem to follow the same cycle as Mn, Fe behaves in a different manner; it was not taken up byMetallogenium at the time of our study.     

Interaction between interstitial water and sediment in two cores of Lac Léman,Switzerland

The pore fluids of the sediments collected at the deepest point of Lac Léman (Switzerland) are supersaturated with respect to vivianite and siderite. In the presence of sulphide, the iron solubility is controlled entirely by the amorphous iron sulphides. As the iron (II) becomes dominant, the formation of siderite occurs and evidence of this, in the solid phase, can be obtained by the use of M?ssbauer spectroscopy and some sequential chemical extractions. The amount of ‘siderite iron’ decreases from about 10% near the sediment surface to a few percent in the lower levels of the sediment (<10 cm). Evidence for vivianite formation could not be obtained even in the lower layers, despite the precautions taken to avoid oxidation. Although the trace metal behaviour in the solid phase is well correlated with the iron and manganese, availability in the pore fluid is dependent on the adsorption on, or co-precipitation with, finely dispersed colloids, which pass through a 0.45 μg filter. Trace metal concentrations in pore fluid were not directly related to total elemental concentrations in the solid phase, and did not reflect cumulative trends associated with anthropogenic enrichment.     

Carbon assimilation during larval development of the marine teleost Leiostomus xanthurus Lacépède

In larval spot, Leiostomus xanthurus Lacépède, ¹⁴C labeled food was rapidly digested and assimilated with up to 20% respired as ¹⁴CO₂ within 10 h from ingestion. Two indices of carbon assimilation, total carbon utilization (range 9.4 to 64.7%) and total carbon absorption (range 67.2 to 99.2%) were calculated from the amount of ¹⁴C ingested, retained, and defecated. Carbon utilization and absorption were not significantly (P > 0.10) related to development as measured by age (7 to 47 days from hatching), length (1.8 to 11.4 mm), or dry weight (0.022 to 4.425 mg).     

漆酶Lac1338的酶学特性测定及定向突变对其酶解染料影响

为了获得表达量高、热稳定性好的漆酶,通过密码子优化合成漆酶基因lac1338、连接到pET-32a(+)载体上并在Escherichia coli BL21(DE3)中表达,获得HIS-Lac1338蛋白。酶学性质测定结果显示,以ABTS为底物时,HIS-Lac1338的比活力高达22.8 U/mg,Km和Vmax分别为567μmol/L和2.8 mmol/(L·min·g);HIS-Lac1338的最适反应温度为55℃,最适pH为6.0;在55℃以下保温2 h能保留50%的酶活性,在pH 4-8范围内孵育4 h仍保留50%以上的活性;HIS-Lac1338对Cu~(2+)抗性强,Ca~(2+)、Na~+、K~+对HIS-Lac1338有促进作用,而Co~(2+)、Fe~(2+)、Hg~(2+)、Ag~+等重金属离子对HIS-Lac1338有抑制作用。易错PCR方法得到的Lac1338的突变酶Lac16与HIS-Lac1338相比,对酸性紫7、溴酚蓝、考马斯亮蓝的降解率分别由10.9%、20%和25%提高到90.5%、67.8%和85%。结果表明,HIS-Lac1338具有较好的温度及pH稳定性,而通过易错PCR技术定向突变获得的突变酶Lac16的染料降解率大大提高。     

Contribution a l'étude du sediment d'un lac oligomésotrophe d'origine volcanique (Lac Pavin,France)

Sediments were collected from a oligomesotrophic lake at different depths: they have been analysed for Ca, Mg, Zn, Na, K, Mn and Fe and organic matter. In the –30 m layer the iron concentration was found to be relatively high, the organic matter concentration low and the C/N ratio very high.

     

Molecular inferences about the genus Hypostomus Lacépède, 1803 (Siluriformes: Loricariidae): a review

This review compiles and discusses the use of genetic markers applied in the study of the fish genus Hypostomus Lacépède, 1803 (Siluriformes: Loricariidae). The database comprises 51 peer-review articles that were published in the last 52 years (1968–2020) and that approach analysis based on different classes of genetic markers. The use of cytogenetic and enzymatic markers was predominantly especially in population studies with the genus Hypostomus, while mitochondrial markers were the majority in phylogenetic studies. Although significant methodological advances have occurred for molecular evaluation, they are still modestly applied to the study of neotropical fish genera, in which Hypostomus is included. New perspectives, especially on integrative approaches, are needed to improve our knowledge of the genetic functionality of fishes.

     

漆酶Lac1338的酶学特性测定及定向突变对其酶解染料影响北大核心CSCD

为了获得表达量高、热稳定性好的漆酶,通过密码子优化合成漆酶基因lac1338、连接到pET-32a(+)载体上并在Escherichia coli BL21(DE3)中表达,获得HIS-Lac1338蛋白。酶学性质测定结果显示,以ABTS为底物时,HIS-Lac1338的比活力高达22.8 U/mg,Km和Vmax分别为567μmol/L和2.8 mmol/(L·min·g);HIS-Lac1338的最适反应温度为55℃,最适pH为6.0;在55℃以下保温2 h能保留50%的酶活性,在pH 4-8范围内孵育4 h仍保留50%以上的活性;HIS-Lac1338对Cu^(2+)抗性强,Ca^(2+)、Na^+、K^+对HIS-Lac1338有促进作用,而Co^(2+)、Fe^(2+)、Hg^(2+)、Ag^+等重金属离子对HIS-Lac1338有抑制作用。易错PCR方法得到的Lac1338的突变酶Lac16与HIS-Lac1338相比,对酸性紫7、溴酚蓝、考马斯亮蓝的降解率分别由10.9%、20%和25%提高到90.5%、67.8%和85%。结果表明,HIS-Lac1338具有较好的温度及pH稳定性,而通过易错PCR技术定向突变获得的突变酶Lac16的染料降解率大大提高。     

Karyotypic diversification in Hypostomus Lacépède, 1803 (Siluriformes, Loricariidae): biogeographical and phylogenetic perspectives

Hypostomus is a species-rich genus of fish with unclear systematics and phylogenetic relationships. Ten species of Hypostomus (H. albopunctatus, H. ancistroides, H. cochliodon, H. commersoni, H. faveolus, H. hermanni, H. aff. paulinus, H. regani, H. strigaticeps and H. topavae) were cytogenetically analyzed through Giemsa staining and silver nitrate impregnation, and the obtained data were correlated to the available biogeographical and phylogenetic analyses for the genus. Although the silver stained nucleolar organizer regions (AgNORs) were found to vary significantly among the species, the diploid numbers could be correlated to the distribution of the species on northern and southern South America river basins. Species with the lower diploid numbers (2n = 64) were associated to northern hydrographic basins and showed a single AgNORs bearing pair. Diploid numbers of 66–68 chromosomes and of 70–84 chromosomes were correlated to two major clades within Hypostomus and southern hydrographic basins, and showed AgNORs varying on number and position. Our results show that cytogenetic data can be correlated to the phylogeny and biogeography of the genus, helping to clarify its complex evolutionary history.     

Comparison of ageing methods and growth rates for largemouth bass,Micropterus salmoides Lacépède,from northern latitudes

Synopsis Ages determined by counts of apparent annuli on scales, sagittae, vertebrae, pectoral fin ray and dorsal fin spine cross sections of largemouth bass from northern populations, which are older and slower growing fish than in the southern parts of its native range, were compared to establish the accuracy of each method. Linear regression techniques indicated strong agreement (r> 0.9) among ages assigned from the examination of scales, sagittae, and vertebrae. The pattern of growth zones on pectoral fin ray and dorsal fm spine cross sections proved too variable for accurate age determination. Limited data suggest that ages greater than 7+ assigned from scales were more likely to underestimate true ages than the other body parts used, although none of these methods gave satisfactory results. Examination of scales from recovered tagged fish, and the similarity between back-calculated lengths of fish through age 7+ to annulus I and observed lengths of juvenile largemouth bass near the end of their first growing season, support the validity of ages determined from scales. Despite a very limited amount of habitat suitable for largemouth bass and severe climatic conditions, growth of this species in Tadenac Lake was similar to growth in other waters north of the Great Lakes. Differences in physical characteristics among these waters does not appear to influence growth rates of largemouth bass, but probably affects production and biomass.     

Differential expression for tissue-specific isozymes in three species of Hypostomus Lacépède, 1803 (Teleostei: Loricariidae)

The expression patterns of 14 enzymatic systems in skeletal muscle, liver and heart tissues of three species of Hypostomus from the Iguaçu River basin (Brazil) were investigated. Although the patterns were similar for the three species, different tissues showed differential expressions, and the data showed that differential tissue expressions of isoperoxidases may be due to preferential combination or association of polypeptide subunits. The detected patterns for SOD isozymes showed that the quaternary structures of these enzymes were in disagreement with the subunit number reported for the majority of other vertebrate groups. Tissue-specific restriction on heterotetramer formation also were described in LDH and MDHP isozymes. Thus, these tissue-specific gene expression character in the species of Hypostomus have the greatest potential to be recognized and applied in systematic studies among species of Hypostomus.     

脓毒症相关急性呼吸窘迫综合征患者血清Lac、D-D、sTM、ESM-1的表达及其临床意义

摘要 目的：探讨脓毒症相关急性呼吸窘迫综合征（ARDS）患者血清乳酸（Lac）、D-二聚体（D-D）、可溶性血栓调节蛋白（sTM）、内皮细胞特异性分子-1（ESM-1）的表达及其临床意义。方法：选取2020年1月～2022年1月徐州医科大学附属医院重症医学科收治的375例脓毒症相关ARDS患者,根据入院时氧合指数（OI）差异分为轻度组（n＝96）、中度组（n＝152）和重度组（n＝127）,比较各组血清Lac、D-D、sTM、ESM-1水平,采用Spearman相关系数分析血清Lac、D-D、sTM、ESM-1与OI的相关性。根据28d内预后将患者分为死亡组（n＝137）和存活组（n＝238）,收集临床资料,采用多因素Logistic回归分析脓毒症相关ARDS患者预后的影响因素。采用受试者工作特征（ROC）曲线分析血清Lac、D-D、sTM、ESM-1对脓毒症相关ARDS患者预后不良的预测价值。结果：轻度组、中度组、重度组血清Lac、D-D、sTM、ESM-1水平依次升高（P＜0.05）。Spearman相关性分析结果显示：脓毒症相关ARDS患者的血清Lac、D-D、sTM、ESM-1水平与OI均呈负相关（P＜0.05）。死亡组年龄大于存活组,脓毒性休克、ICU停留时间≥10 d、机械通气时间≥3 d比例以及SOFA评分、血清Lac、D-D、sTM、ESM-1水平均高于存活组,OI低于存活组（P＜0.05）。多因素Logistic回归分析显示：脓毒性休克、SOFA评分≥13分、血清Lac、D-D、sTM、ESM-1水平较高是脓毒症相关ARDS患者预后不良的危险因素,而OI较高是预后不良的保护因素（P＜0.05）。ROC曲线分析结果显示：血清Lac、D-D、sTM、ESM-1联合检测预测脓毒症相关ARDS患者预后不良的AUC为0.907,明显高于各指标单独检测的0.785、0.747、0.796、0.789。结论：血清Lac、D-D、sTM、ESM-1水平升高与脓毒症相关ARDS病情严重程度和预后不良密切相关,对患者预后有一定预测价值。     

A study of spatial and temporal variability in the adenine nucleotides of lake phytoplankton during a diel cycle (Lac Pavin — France)

A study of diel variations in the adenine nucleotide concentrations of natural phytoplanktonic populations led to the following conclusions:There are significant diel variations in cellular ATP concentrations. However, this temporal variability is not as great as the spatial variability between the various sampling depths, characterized by different phytoplankton populations. Within a globally stable pool of adenine nucleotides, diel variations inside the algal cell mainly take place at the expense of ATP and AMP, while ADP concentrations remain relatively stable.The statistical relationship between algal biomass obtained from cell counts and from adenine nucleotides also confirms maximum stability for ADP and (ATP + ADP + AMP). The diel variations in adenine nucleotide cell concentrations gradually become smaller with depth. This probably reflects the importance of light in causing such variability.     

Comparative studies of bile salts. 5α-Chimaerol, a new bile alcohol from the white sucker Catostomus commersoni Lacépède

1. G.l.c. examination of bile alcohols prepared from the sucker Catostomus commersoni Lacépède (family Catostomidae) showed that although 5α-cyprinol (5α-cholestane-3α,7α,12α,26,27-pentol) was a minor constituent, the principal bile alcohol was an undescribed substance, probably present in the bile as the C-26 sulphate ester, whose i.r., n.m.r. and mass spectra agreed with the structure 5α-cholestane-3α,7α,12α,24,26-pentol. 2. M_D studies suggest that this 5α-chimaerol is the 24(+), 25S enantiomer and that 5β-chimaerol (chimaerol) from Chimaera monstrosa bile also has the 24(+), 25S configuration. These findings imply that bile alcohol biosynthesis in suckers and chimaeras includes stereospecific oxidation of cholesterol at C-26. 3. C. commersoni bile acids (present in minor amounts) probably consist largely of 3α,7α,12α-trihydroxy-5α-cholan-24-oic acid (allocholic acid). 4. 5α-Chimaerol sulphate and 5α-cyprinol sulphate are probably biochemically equivalent as bile salts, and can be considered as arising by parallel evolution.     

A New Species of Trimeresurus Lacépède, 1804(Squamata: Viperidae) from Southwestern China,Vietnam, Thailand and Myanmar

The pit vipers of the genus Trimeresurus Lacépède, 1804 is one of the largest groups of Asian snakes, distributed from India to China and Indonesia. Recent surveys in Jiangcheng and Simao, Yunnan Province, China resulted in a new species previously allocated to T. albolabris. Combining morphological and molecular data, we describe it as Trimeresurus guoi sp. nov. The new species morphologically differs from T. albolabris in the yellow green ventral color; an indistinct ventrolateral line; the a bsence of a postocular stripe; the firebrickred iris; a dark red stripe on dorsal tail; hemipenes with relatively weak sparse papillae, reaching 23 rd subcaudal when unextruded. Molecularly, the new species forms a clearly divergent lineage(BPP 1.00/UFB 100). Uncorrected pairwise distances of mitochondrial gene Cyt b between the new species and other known species of the subgenus Trimeresurus range from 0.052(T. albolabris) to 0.071(T. insularis).