Overestimated accuracy of circular dichroism in determining protein secondary structure

Circular dichroism (CD) is a spectroscopic technique widely used for estimating protein secondary structures in aqueous solution, but its accuracy has been doubted in recent work. In the present paper, the contents of nine globular proteins with known secondary structures were determined by CD spectroscopy and Fourier transform infrared spectroscopy (FTIR) in aqueous solution. A large deviation was found between the CD spectra and X-ray data, even when the experimental conditions were optimized. The content determined by FTIR was in good agreement with the X-ray crystallography data. Therefore, CD spectra are not recommended for directly calculating the content of a protein’s secondary structure.     

Roles of sulfate adsorption and base cation supply in controlling the chemical response of streams of western Virginia to reduced acid deposition

Micro-organisms are vital for the functioning of all food webs and are the major drivers of the global biogeochemical cycles. The microbial community compositions and physicochemical conditions of the different water masses in the North Sea, a biologically productive sea on the northwestern European continental shelf, were studied during two summer cruises, in order to provide detailed baseline data for this region and examine its microbial biogeography. For each cruise the stations were clustered according to their physicochemical characteristics and their microbial community composition. The largest cluster, which covered most of the central and northern North Sea, consisted of stations that were characterized by a thermally stratified water column and had low chlorophyll a autofluorescence and generally low microbial abundances. The second main cluster contained stations that were dominated by picoeukaryotes and showed the influence of influxes of North Atlantic water via the English Channel and south of the Shetland Islands. The third main cluster was formed by stations that were dominated by cyanobacteria and nanoeukaryotes in the reduced salinity Norwegian Coastal and Skagerrak waters, while the fourth cluster represented the German Bight, a region with strong riverine input, high nutrient concentrations, and consequently high heterotrophic bacterial and viral abundances. Despite the complex and dynamic hydrographic nature of the North Sea, the consistent distinctions in microbiology between these different hydrographic regions during both cruises illustrate the strong links between the microbial community and its environment, as well as the possibility to use microorganisms for long-term monitoring of environmental change.     

Apelin-36, a potent peptide,protects against ischemic brain injury by activating the PI3K/Akt pathway

Apelin is an endogenous ligand of G protein-coupled receptor-apelin and angiotensin-1-like receptor (APJ). The biological effects of apelin–APJ system are reported in multiple systems including cardiovascular, endocrinal, and gastrointestinal system. Previous studies had shown that apelin-13 is a potential protective agent on cardiac ischemia; however, the role of apelin in the central nervous system remained unknown. In this study, we investigated therapeutic effects of apelin-36, a long form of apelin, in ischemic brain injury models. We found that apelin-36 reduced cerebral infarct volume in the middle cerebral artery occlusion (MCAO) model and the neonatal hypoxic/ischemic (H/I) injury model. Apelin-36 improved neurological deficits in the MCAO model and promoted long-term functional recovery after H/I brain injury. We further explored the protective mechanisms of apelin-36 on H/I brain injury. We clearly demonstrated that apelin-36 significantly reduced the levels of cleaved caspase-3 and Bax, two well-established apoptotic markers after H/I injury, indicating the anti-apoptotic activity of apelin-36 in ischemic injury. Since apelin-36 increased the level of phosphorylated Akt after H/I injury, we treated neonates with a specific PI3K inhibitor LY294002. We found that LY294002 decreased the phosphorylated Akt level and attenuated protective effects of apelin-36 on apoptosis. These suggested that the PI3K/Akt pathway was at least in part involved in the anti-apoptotic mechanisms of apelin-36. Our findings demonstrated that apelin-36 was a promising therapeutic agent on the treatment of ischemic brain injury.     

Percutaneous comprehensive cryoablation for metastatic esophageal cancer after failure of radical surgery

Esophageal cancer is common in China. There is a lack of treatment strategies for metastatic esophageal cancer (MEC) after radical surgery on the primary tumor. Cryoablation is an attractive option because tumor necrosis can be safely induced in a minimally invasive manner. This study assessed its therapeutic effect in MEC after failure of radical surgery. One hundred and forty patients met the inclusion criteria from May, 2003 to March, 2011. Comprehensive cryotherapy of multiple metastases was performed on 105 patients; 35 received chemotherapy. No severe complications occurred during or after cryoablation. Overall survival (OS) was assessed according to therapeutic protocol, pathologic type, treatment timing and number of procedures. The OS of patients who received comprehensive cryoablation (44 ± 20 months) was significantly longer than that of those who underwent chemotherapy (23 ± 24 months; P = 0.0006). In the cryotherapy group, the OS for squamous cell carcinoma (45 ± 19 months) was longer than that for adenocarcinoma (33 ± 18 months; P = 0.0435); the OS for timely cryoablation (46 ± 19 months) was longer than that for delayed cryoablation (33 ± 20 months; P = 0.0193); the OS for multiple cryoablation (50 ± 17 months) was longer than that for single cryoablation (37 ± 20 months; P = 0.0172); and the OS for cryo-immunotherapy (56 ± 17 months) was longer than that for cryoablation alone (39 ± 19 months; P = 0.0011). Thus, comprehensive cryotherapy may have advantages over chemotherapy in the treatment of MEC and, in patients with squamous cell carcinoma, supplementary immunotherapy and timely and multiple cryoablation may be associated with a better prognosis.     

Submillisecond Elastic Recoil Reveals Molecular Origins of Fibrin Fiber Mechanics

Fibrin fibers form the structural scaffold of blood clots. Thus, their mechanical properties are of central importance to understanding hemostasis and thrombotic disease. Recent studies have revealed that fibrin fibers are elastomeric despite their high degree of molecular ordering. These results have inspired a variety of molecular models for fibrin’s elasticity, ranging from reversible protein unfolding to rubber-like elasticity. An important property that has not been explored is the timescale of elastic recoil, a parameter that is critical for fibrin’s mechanical function and places a temporal constraint on molecular models of fiber elasticity. Using high-frame-rate imaging and atomic force microscopy-based nanomanipulation, we measured the recoil dynamics of individual fibrin fibers and found that the recoil was orders of magnitude faster than anticipated from models involving protein refolding. We also performed steered discrete molecular-dynamics simulations to investigate the molecular origins of the observed recoil. Our results point to the unstructured αC regions of the otherwise structured fibrin molecule as being responsible for the elastic recoil of the fibers.     

Biomimetic injectable HUVEC‐adipocytes/collagen/alginate microsphere co‐cultures for adipose tissue engineering

Engineering adipose tissue that has the ability to engraft and establish a vascular supply is a laudable goal that has broad clinical relevance, particularly for tissue reconstruction. In this article, we developed novel microtissues from surface‐coated adipocyte/collagen/alginate microspheres and human umbilical vein endothelial cells (HUVECs) co‐cultures that resembled the components and structure of natural adipose tissue. Firstly, collagen/alginate hydrogel microspheres embedded with viable adipocytes were obtained to mimic fat lobules. Secondly, collagen fibrils were allowed to self‐assemble on the surface of the microspheres to mimic collagen fibrils surrounding the fat lobules in the natural adipose tissue and facilitate HUVEC attachment and co‐cultures formation. Thirdly, the channels formed by the gap among the microspheres served as the room for in vitro prevascularization and in vivo blood vessel development. The endothelial cell layer outside the microspheres was a starting point of rapid vascular ingrowth. Adipose tissue formation was analyzed for 12 weeks at 4‐week intervals by subcutaneous injection into the head of node mice. The vasculature in the regenerated tissue showed functional anastomosis with host blood vessels. Long‐term stability of volume and weight of the injection was observed, indicating that the vasculature formed within the constructs benefited the formation, maturity, and maintenance of adipose tissue. This study provides a microsurgical method for adipose regeneration and construction of biomimetic model for drug screening studies. Biotechnol. Bioeng. 2013; 110: 1430–1443. © 2012 Wiley Periodicals, Inc.     

A biofilm model to understand the onset of sulfate reduction in denitrifying membrane biofilm reactors

This work presents a multispecies biofilm model that describes the co‐existence of nitrate‐ and sulfate‐reducing bacteria in the H₂‐based membrane biofilm reactor (MBfR). The new model adapts the framework of a biofilm model for simultaneous nitrate and perchlorate removal by considering the unique metabolic and physiological characteristics of autotrophic sulfate‐reducing bacteria that use H₂ as their electron donor. To evaluate the model, the simulated effluent H₂, UAP (substrate‐utilization‐associated products), and BAP (biomass‐associated products) concentrations are compared to experimental results, and the simulated biomass distributions are compared to real‐time quantitative polymerase chain reaction (qPCR) data in the experiments for parameter optimization. Model outputs and experimental results match for all major trends and explain when sulfate reduction does or does not occur in parallel with denitrification. The onset of sulfate reduction occurs only when the nitrate concentration at the fiber's outer surface is low enough so that the growth rate of the denitrifying bacteria is equal to that of the sulfate‐reducing bacteria. An example shows how to use the model to design an MBfR that achieves satisfactory nitrate reduction, but suppresses sulfate reduction. Biotechnol. Bioeng. 2013; 110: 763–772. © 2012 Wiley Periodicals, Inc.     

钝齿棒杆菌argR基因缺失株构建及其缺失对精氨酸生物合成途径相关基因转录水平的影响

[目的]钝齿棒杆菌AS 1.542中argR基因编码的蛋白ArgR在精氨酸生物合成途径中扮演负调控的角色,但其对相关基因在转录水平的影响还未见报道.因此,本课题组构建了钝齿棒杆菌argR基因缺失株,并在转录水平上比较野生株与缺失株精氨酸生物合成途径相关基因的变化.[方法]采用无痕敲除的方法构建了钝齿棒杆菌argR基因缺失株,并采用荧光定量PCR方法分析缺失株和野生株精氨酸生物合成途径相关基因在转录水平的变化.[结果]利用pK18mobsacB质粒中蔗糖致死基因sacB反向筛选标记及PCR方法成功筛选到钝齿棒杆菌argR基因缺失株;荧光定量PCR结果表明,argR基因缺失株精氨酸生物合成途径中相关基因在转录水平获得大量提高,平均约上调162.13倍.[结论]钝齿棒杆菌精氨酸生物合成途径的相关基因受负调控蛋白ArgR的显著调控,但其基因的敲除并没有引起精氨酸产量发生明显的变化.     

Pip介导铜绿假单胞菌吩嗪基因簇phz2的表达

[目的]为了确定铜绿假单胞菌调控因子Pip对两个不同吩嗪合成基因簇(phz1和phz2)的具体调控方式与可能的调控机制.[方法]根据基因比对结果,采用同源重组技术构建Pip调控因子缺失突变株PA-PG以及克隆ip基因作互补分析;再以已构建的吩嗪基因簇缺失突变株PA-Z1G和PA-Z2K为受体菌,构建突变株PA-PD-Z1G和PA-PG-Z2K,测定并比较野生株及相关突变株的吩嗪-1-羧酸和绿脓菌素的合成量,推定Pip对两个不同吩嗪合成基因簇的调控方式.[结果]在GA培养基中,突变株PA-PG的吩嗪-1-羧酸和绿脓菌素都比野生型明显减少;互补分析显示,突变株PA-PG的吩嗪-1-羧酸和绿脓菌素都显著提高并恢复到野生株PAO1水平;突变株PA-Z1G的吩嗪-1-羧酸和绿脓菌素合成量因Pip缺失而显著减少;而突变株PA-Z2K的吩嗪-1-羧酸和绿脓菌素合成量在Pip缺失后仍保持不变.[结论]初步推定,转录调控因子Pip对铜绿假单胞菌吩嗪合成代谢的确具有促进作用;Pip通过正向调控吩嗪基因簇phz2的合成功能实现对吩嗪合成代谢的调控.