1H NMR identification of a beta-sheet structure and description of folding topology in putidaredoxin

Putidaredoxin (Pdx), a 106-residue globular protein consisting of a single polypeptide chain and a [2Fe-2S] cluster, is the physiological reductant of P-450cam, which in turn catalyzes the monohydroxylation of camphor by molecular oxygen. No crystal structure has been obtained for Pdx or for any closely homologous protein. The application of two-dimensional 1H NMR methods to the problem of structure determination in Pdx is reported. A beta-sheet consisting of five short strands and one beta-turn has been identified from distinctive nuclear Overhauser effect patterns. All of the backbone resonances and a majority of the side-chain resonances corresponding to protons in the beta-sheet have been assigned sequence specifically. The sheet contains one parallel and three antiparallel strand orientations. Hydrophobic side chains in the beta-sheet face primarily toward the protein interior, except for a group of three valine side chains that are apparently solvent exposed. The potential significance of this "hydrophobic patch" in terms of biological activity is discussed. The folding topology, as determined by the constraints of the beta-sheet, is compared with that of other [2Fe-2S] proteins for which folding topologies are known.     

Relaxation dynamics of the gel to liquid-crystalline transition of phosphatidylcholine bilayers. Effects of chainlength and vesicle size.

The relaxation kinetics of the gel to liquid-crystalline transition of five phosphatidylcholine (DC14PC to DC18PC) bilayer dispersions have been investigated using volume perturbation calorimetry, a steady-state technique which subjects a sample to sinusoidal changes in volume. Temperature and pressure responses to the volume perturbation are measured to monitor the relaxation to a new equilibrium position. The amplitude demodulation and phase shift of these observables are analyzed with respect to the perturbation frequency to yield relaxation times and amplitudes. In the limit of low perturbation frequency, the temperature and pressure responses are proportional to the equilibrium excess heat capacity and bulk modulus, respectively. At all temperatures, the thermal response data are consistent with a single primary relaxation process of the lipid. The less accurate bulk modulus data exhibit two relaxation times, but it is not clear whether they reflect lipid processes or are characteristic of the instrument. The observed thermal relaxation behavior of all multilamellar vesicles are quantitatively similar. The relaxation times vary from approximately 50 ms to 4 s, with a pronounced maximum at a temperature just greater than Tm, the temperature of the excess heat capacity maximum. Large unilamellar vesicles also exhibit a single relaxation process, but without a pronounced maximum in the relaxation time. Their relaxation time is approximately 80 ms over most of the transition range.     

Effect of cellulases on spontaneous fusion of maize protoplasts

Summary The effect of protoplast-isolating enzymes on spontaneous fusion of maize protoplasts (Zea mays L. cv. Black Mexican Sweet) was investigated using a convenient ethidium bromide nuclear staining procedure. After 2–2.5 hour digestion in an enzyme solution containing 1% Cellulysin, 0.5% Rhozyme, and 0.02% Pectolyase Y-23, 50–75% of the protoplasts contained multiple nuclei. The cellulase Cellulysin was identified as the factor causing the spontaneous protoplast fusion; when Cellulysin was replaced by CELF cellulase, most protoplasts were uninucleate. Calcium and other components in the enzyme solution did not affect spontaneous fusion. Cellulysin also increased the percentage of multinucleate protoplasts from rice and asparagus suspensions. Presence of multiple nuclei might affect genetic manipulations involving protoplasts.     

低温胁迫下凤眼莲叶片的适应——脱落酸和可溶性蛋白质含量升高

本文报道低温胁迫下风眼莲叶片脱落酸(ABA)、可溶性蛋白质和水势的测定结果。低温胁迫时脱落酸和可溶性蛋白质含量远高于对照,(前者含量最高可达对照的4倍,后者可达到对照的2.75倍),而且脱落酸和蛋白质含量随温度降低而升高。蛋白质的生物合成抑制剂亚胺环己酮证明,可溶性蛋白质含量升高,原因是有部分是新合成的。在各种低温处理下获得了几乎相同于对照的叶片水势。我们推测:低温胁迫下,脱落酸水平的相应变化不是由于低温诱导水分胁迫所致,而是低温胁迫本身诱导。     

荒漠结皮层藓类植物死亡对表层土壤水分蒸发和入渗的影响

水分是荒漠植物生长最主要的限制因子,藓类结皮作为荒漠土壤表层重要覆被物,对土壤水分蒸发入渗具有重要影响。研究表明,在全球气候变化背景下,不确定的降水格局变化导致结皮层藓类植物出现集群死亡现象,但这一过程对荒漠地表土壤水分蒸发与入渗过程的影响及其机理尚不清楚。以古尔班通古特沙漠齿肋赤藓结皮为研究对象,利用便携式渗透计和蒸发仪,研究了结皮层藓类植物死亡对土壤水分蒸发与入渗的影响。结果表明,与裸沙相比,藓类结皮的存在显著抑制了水分入渗,而藓类植物死亡的结皮层抑制作用最大,其初渗速率、稳渗速率和累积入渗量分别是活藓类结皮的39.89%、85.91%及64.48%,仅为裸沙的5.96%、13.13%及20.42%。在水分蒸发初期,裸沙的水分蒸发速率明显高于活藓类结皮和藓类植物死亡的结皮层,但藓类植物死亡的结皮层维持相对稳定的蒸发速率的时间长于裸沙和活藓类结皮,这也导致最终累计蒸发量以藓类植物死亡的结皮层最高、裸沙最低。可见,荒漠生物土壤结皮中藓类植物死亡会明显减少土壤水分入渗、增大水分蒸发,进一步影响荒漠表层土壤水分格局,从而影响生物土壤结皮与维管植物的水分利用关系。     

基于生态系统评价的山水林田湖草生态保护与修复体系构建研究——以乌梁素海流域为例

山水林田湖草系统保护与修复工程实施的重要目标是维护和提升区域生态系统服务。从乌梁素海流域山水林田湖草的生态现状与功能出发,对乌梁素海流域水土流失、土地沙化等生态敏感性及土壤保持、水源涵养、生物多样性等生态功能重要性进行系统评价,形成基于生态敏感性和生态功能重要性相结合的空间格局评价结果。以维护和提升人类福祉所需的重要生态系统服务为目标,以乌梁素海流域生态敏感性和生态功能重要性相结合的空间格局评价结果为基础,制定了“一中心、二重点、六要素、七工程”的乌梁素海山水林田湖草生态保护与修复体系,并基于此将乌梁素海流域生态保护修复分为6个主要治理区域,形成“四区、一带、一网”的生态安全格局,通过具体工程实施,流域生态环境质量和生态服务能力将取得明显提升,防风固沙能力有效增强,生物多样性持续改善,水环境质量稳定达标,生态系统的稳定性明显加强。通过乌梁素海流域的分析案例为流域山水林田湖草生态保护与修复关键区域的识别提供了定量分析方法,为流域尺度构建生态安全格局、实现山水林田湖草系统保护和修复提供思路和途径。     

基于形态学空间格局分析和最小累积阻力模型的城市生态安全格局构建——以厦门市为例

城市化进程的快速发展加剧了生态系统的退化。如何扭转生态系统的退化,同时满足人类日益增长的生态系统服务需求,成为当前的一个研究热点。生态安全格局的构建在一定程度上可平衡城市发展与生态环境保护之间的关系,对于保障区域生态安全、提升生态系统功能具有重大意义。以厦门市为例,基于“生态源地识别—阻力面构建—生态廊道提取”的基本框架构建陆域生态安全格局。结合生态系统服务重要性评价和形态学空间格局分析识别生态源地,该方法兼顾了生态结构和功能,使得所识别的生态源地更具全面性。选取土地利用类型、高程和坡度构建生态综合阻力面,并用人类居住合成指数修正生态综合阻力面,以减少主观赋值的影响,识别各土地利用类型内部的差异,使生态阻力面的构建更加合理。在此基础上通过最小累积阻力模型提取生态廊道,利用重力模型量化潜在生态廊道的相对重要性,并根据重力模型结果划分重要性等级。研究结果表明,厦门市的生态安全格局由14个生态源地、21条生态廊道、15个生态节点及若干个踏脚石所组成。生态源地主要集中在研究区的西部和北部,以林地和草地为主,面积合计为558.64 km²。生态廊道长约159.40 km,其...     

The novel type II toxin–antitoxin PacTA modulates Pseudomonas aeruginosa iron homeostasis by obstructing the DNA-binding activity of Fur

Type II toxin–antitoxin (TA) systems are widely distributed in bacterial and archaeal genomes and are involved in diverse critical cellular functions such as defense against phages, biofilm formation, persistence, and virulence. GCN5-related N-acetyltransferase (GNAT) toxin, with an acetyltransferase activity-dependent mechanism of translation inhibition, represents a relatively new and expanding family of type II TA toxins. We here describe a group of GNAT-Xre TA modules widely distributed among Pseudomonas species. We investigated PacTA (one of its members encoded by PA3270/PA3269) from Pseudomonas aeruginosa and demonstrated that the PacT toxin positively regulates iron acquisition in P. aeruginosa. Notably, other than arresting translation through acetylating aminoacyl-tRNAs, PacT can directly bind to Fur, a key ferric uptake regulator, to attenuate its DNA-binding affinity and thus permit the expression of downstream iron-acquisition-related genes. We further showed that the expression of the pacTA locus is upregulated in response to iron starvation and the absence of PacT causes biofilm formation defect, thereby attenuating pathogenesis. Overall, these findings reveal a novel regulatory mechanism of GNAT toxin that controls iron-uptake-related genes and contributes to bacterial virulence.