Covalent binding of α-chymotrypsin on the magnetic nanogels covered by amino groups

A new aminated carrier—magnetic nanogels covered by amino groups, was obtained by Hoffman degradation of polyacrylamide-coated Fe₃O₄ nanoparticles prepared by photochemical polymerization. α-Chymotrypsin (CT) was covalently bound to the magnetic nanogels by use of 1-ethyl-3-(3-dimethylaminepropyl) carbodiimide and N-hydroxysuccinimide at room temperature. Immobilization time, pH value of the reaction mixture and proportion of CT to the magnetic nanogels were investigated to obtain the optimum condition for CT immobilization. The maximal specific activity of the bound CT was determined to be 0.93 U/(mg min), 59.3% of free counterpart. The maximal binding capacity was measured to be 102 mg enzyme/g nanogel. Furthermore, the bound CT exhibited good thermal stability, storage stability and reusability.     

基于比较基因组学的高地芽孢杆菌6ww6分子标识筛选及其快速鉴定

【目的】高地芽孢杆菌(Bacillus altitudinis) 6ww6是一株根际耐盐促生菌,可以作为微生物肥料的优选菌种。为了加强菌种知识产权保护,建立菌株快速检测方法。【方法】本研究通过基因组比对分析、TBLASTn检索、NR库检索验证并剔除有其他同源的结果序列,最终得到菌株6ww6的孤儿基因,设计引物进行PCR鉴定。【结果】筛选出5个特异性基因J939_13195、J9319_05960、J9319_13355、J9319_05965和J9319_13350。引物5960F和5960R可以单一性扩增出菌株6ww6的目的基因J9319_05960,确定其为菌株6ww6的特异性分子标识。【结论】本研究确定了菌株6ww6的唯一性身份编码,建立了以比较基因组学和孤儿基因为基础的菌株水平上的鉴定方法。该方法具有快速、精确、能鉴定到菌株水平的优点,对于有价值微生物的知识产权保护提供了有力的抓手。     

西南三江(金沙江、澜沧江和怒江)流域化学风化过程

选择中国西南三江流域作为研究对象,结合水化学及溶解性无机碳碳同位素,借助正演模型,分析河水溶解性组分来源及混入比例,说明硫酸参与区域碳酸盐矿物风化过程及CO2净释放量。结果表明:(1)西南三江流域河水受复杂岩性控制,金沙江河水以Na-Ca-Cl-HCO3为主,与流域上游蒸发盐矿物溶解有关,澜沧江和怒江则以Ca-HCO3为主,显示碳酸盐和硅酸盐矿物溶解的影响;(2)蒸发盐矿物溶解是金沙江干流河水阳离子主要来源,其贡献均值为52%,对澜沧江和怒江河水贡献较小,平均值分别为11%和2%。碳酸盐矿物溶解是澜沧江和怒江干流河水阳离子主要来源,其贡献均值分别为70%和78%,对金沙江干流河水的贡献较小,平均为38%,但对金沙江支流河水的贡献较大,平均为74%。硅酸盐风化产物对金沙江、澜沧江和怒江干流的平均贡献分别为8%、16%和15%,对金沙江支流河水的贡献为15%;(3)金沙江、澜沧江和怒江流域硅酸盐矿物风化速率分别为1.39、3.27和4.27 t·km-2·a-1,CO2消耗速率分别为0.34×105、1.18×105和1.40×105mol·km-2·a-1,碳酸盐矿物风化速率分别为16.93、33.13和33.54 t·km-2·a-1,CO2消耗速率分别为1.40×105、2.47×105和2.22×105mol·km-2·a-1;(4)西南三江流域河水硫酸盐主要来源于硫化物矿物氧化,硫酸参与碳酸盐矿物风化净释放CO2量分别为0.73×105、3.01×105和4.27×105mol·km-2·a-1,高于中国贵州地区,应当在研究区域碳循环以及全球气候变化过程中引起重视。     

Biomimetic repeat protein derived from Xenopus tropicalis for fibrous scaffold fabrication

Collagen, silk, and elastin are the fibrous proteins consist of representative amino acid repeats. Because these proteins exhibited distinguishing mechanical properties, they have been utilized in diverse applications, such as fiber‐based sensors, filtration membranes, supporting materials, and tissue engineering scaffolds. Despite their infinite prevalence and potential, most studies have only focused on a few repeat proteins. In this work, the hypothetical protein with a repeat motif derived from the frog Xenopus tropicalis was obtained and characterized for its potential as a novel protein‐based material. The codon‐optimized recombinant frog repeat protein, referred to as ‘xetro’, was produced at a high rate in a bacterial system, and an acid extraction‐based purified xetro protein was successfully fabricated into microfibers and nanofibers using wet spinning and electrospinning, respectively. Specifically, the wet‐spun xetro microfibers demonstrated about 2‐ and 1.5‐fold higher tensile strength compared with synthetic polymer polylactic acid and cross‐linked collagen, respectively. In addition, the wet‐spun xetro microfibers showed about sevenfold greater stiffness than collagen. Therefore, the mass production potential and greater mechanical properties of the xetro fiber may result in these fibers becoming a new promising fiber‐based material for biomedical engineering. © 2015 Wiley Periodicals, Inc. Biopolymers 103: 659–664, 2015.     

The Atomic Structure of the HIV-1 gp41 Transmembrane Domain and Its Connection to the Immunogenic Membrane-proximal External Region

The membrane-proximal external region (MPER) C-terminal segment and the transmembrane domain (TMD) of gp41 are involved in HIV-1 envelope glycoprotein-mediated fusion and modulation of immune responses during viral infection. However, the atomic structure of this functional region remains unsolved. Here, based on the high resolution NMR data obtained for peptides spanning the C-terminal segment of MPER and the TMD, we report two main findings: (i) the conformational variability of the TMD helix at a membrane-buried position; and (ii) the existence of an uninterrupted α-helix spanning MPER and the N-terminal region of the TMD. Thus, our structural data provide evidence for the bipartite organization of TMD predicted by previous molecular dynamics simulations and functional studies, but they do not support the breaking of the helix at Lys-683, as was suggested by some models to mark the initiation of the TMD anchor. Antibody binding energetics examined with isothermal titration calorimetry and humoral responses elicited in rabbits by peptide-based vaccines further support the relevance of a continuous MPER-TMD helix for immune recognition. We conclude that the transmembrane anchor of HIV-1 envelope is composed of two distinct subdomains: 1) an immunogenic helix at the N terminus also involved in promoting membrane fusion; and 2) an immunosuppressive helix at the C terminus, which might also contribute to the late stages of the fusion process. The unprecedented high resolution structural data reported here may guide future vaccine and inhibitor developments.     

Structure-Function Analysis of CCL28 in the Development of Post-viral Asthma

CCL28 is a human chemokine constitutively expressed by epithelial cells in diverse mucosal tissues and is known to attract a variety of immune cell types including T-cell subsets and eosinophils. Elevated levels of CCL28 have been found in the airways of individuals with asthma, and previous studies have indicated that CCL28 plays a vital role in the acute development of post-viral asthma. Our study builds on this, demonstrating that CCL28 is also important in the chronic post-viral asthma phenotype. In the absence of a viral infection, we also demonstrate that CCL28 is both necessary and sufficient for induction of asthma pathology. Additionally, we present the first effort aimed at elucidating the structural features of CCL28. Chemokines are defined by a conserved tertiary structure composed of a three-stranded β-sheet and a C-terminal α-helix constrained by two disulfide bonds. In addition to the four disulfide bond-forming cysteine residues that define the traditional chemokine fold, CCL28 possesses two additional cysteine residues that form a third disulfide bond. If all disulfide bonds are disrupted, recombinant human CCL28 is no longer able to drive mouse CD4+ T-cell chemotaxis or in vivo airway hyper-reactivity, indicating that the conserved chemokine fold is necessary for its biologic activity. Due to the intimate relationship between CCL28 and asthma pathology, it is clear that CCL28 presents a novel target for the development of alternative asthma therapeutics.     

A Mechanism of Global Shape-dependent Recognition and Phosphorylation of Filamin by Protein Kinase A

Protein phosphorylation mediates essentially all aspects of cellular life. In humans, this is achieved by ∼500 kinases, each recognizing a specific consensus motif (CM) in the substrates. The majority of CMs are surface-exposed and are thought to be accessible to kinases for phosphorylation. Here we investigated the archetypical protein kinase A (PKA)-mediated phosphorylation of filamin, a major cytoskeletal protein that can adopt an autoinhibited conformation. Surprisingly, autoinhibited filamin is refractory to phosphorylation by PKA on a known Ser²¹⁵² site despite its CM being exposed and the corresponding isolated peptide being readily phosphorylated. Structural analysis revealed that although the CM fits into the PKA active site its surrounding regions sterically clash with the kinase. However, upon ligand binding, filamin undergoes a conformational adjustment, allowing rapid phosphorylation on Ser²¹⁵². These data uncover a novel ligand-induced conformational switch to trigger filamin phosphorylation. They further suggest a substrate shape-dependent filtering mechanism that channels specific exposed CM/kinase recognition in diverse signaling responses.     

Zinc Inhibits Hedgehog Autoprocessing: LINKING ZINC DEFICIENCY WITH HEDGEHOG ACTIVATION*

Zinc is an essential trace element with wide-ranging biological functions, whereas the Hedgehog (Hh) signaling pathway plays crucial roles in both development and disease. Here we show that there is a mechanistic link between zinc and Hh signaling. The upstream activator of Hh signaling, the Hh ligand, originates from Hh autoprocessing, which converts the Hh precursor protein to the Hh ligand. In an in vitro Hh autoprocessing assay we show that zinc inhibits Hh autoprocessing with a K_i of 2 μm. We then demonstrate that zinc inhibits Hh autoprocessing in a cellular environment with experiments in primary rat astrocyte culture. Solution NMR reveals that zinc binds the active site residues of the Hh autoprocessing domain to inhibit autoprocessing, and isothermal titration calorimetry provided the thermodynamics of the binding. In normal physiology, zinc likely acts as a negative regulator of Hh autoprocessing and inhibits the generation of Hh ligand and Hh signaling. In many diseases, zinc deficiency and elevated level of Hh ligand co-exist, including prostate cancer, lung cancer, ovarian cancer, and autism. Our data suggest a causal relationship between zinc deficiency and the overproduction of Hh ligand.     

Synthesis and characterisation of a novel tubulin-directed DO3A-colchicine conjugate with potential theranostic features

Colchicine is a known tubulin binding agent enabling necrosis in tumors. A novel tubulin-directed DO3A-colchicine conjugate and its Gd(III) complex were prepared from N-deacetylcolchicine, coupling alkaloid and polyaza-alicyclic functions via a peptide coupling methodology. The longitudinal proton relaxivity of the Gd(III) complex in water at 4.7 T is 2.86 mM⁻¹ s⁻¹ and a similar efficacy as colchicine towards ovarian carcinoma cells in vitro.     

Property and wealth inequality as cultural niche construction

In contrast to other approaches, evolutionary perspectives on understanding the power and wealth inequalities in human societies view wealth and power not as ends in themselves but as proximate goals that contribute to the ultimate Darwinian goal of achieving reproductive success. The most successful means of achieving it in specific times and places depend on local conditions and these have changed in the course of human history, to such an extent that strategies focused on the maintenance and increase of wealth can even be more successful in reproductive terms than strategies directed at maximizing reproductive success in the short term. This paper argues that a major factor leading to such changes is a shift in the nature of inter-generational wealth transfers from relatively intangible to material property resources and the opportunities these provided for massively increased inequality. This shift can be seen as a process of niche construction related to the increasing importance of fixed and defensible resources in many societies after the end of the last Ice Age. It is suggested that, despite problems of inference, the evidence of the archaeological record can be used to throw light on these processes in specific places and times.