Myotoxic activity of a Gln-49 phospholipase A2 from Agkistrodon blomhoffii ussurensis snake venom

A novel myotoxic protein phospholipase A₂ (PLA₂), denoted as Gln49-PLA₂, has been isolated from snake venom of Agkistrodon blomhoffii ussurensis, which has weak lethal effect and apparent anticoagulant activity, but lacks the PLA₂ and hemorrhagenic activity. Gln49-PLA₂ obviously increases of plasma creatine-kinase (CK) upon intramuscular injection in mice, suggesting that it may induce a dose-dependent myonecrosis. Histological studies also reveal morphological changes in mouse skeletal muscles, including extensive myonecrosis, hemorrhage and neutrophil infiltration in the treated animals. The myotoxic ability induced by Gln49-PLA₂ can be partially inhibited by heparin.     

Myotoxic activity of a Gln-49 phospholipase A2 from Agkistrodon blomhoffii ussurensis snake venom

A novel myotoxic protein phospholipase A₂ (PLA₂), denoted as Gln49-PLA₂, has been isolated from snake venom of Agkistrodon blomhoffii ussurensis, which has weak lethal effect and apparent anticoagulant activity, but lacks the PLA₂ and hemorrhagenic activity. Gln49-PLA₂ obviously increases of plasma creatine-kinase (CK) upon intramuscular injection in mice, suggesting that it may induce a dose-dependent myonecrosis. Histological studies also reveal morphological changes in mouse skeletal muscles, including extensive myonecrosis, hemorrhage and neutrophil infiltration in the treated animals. The myotoxic ability induced by Gln49-PLA₂ can be partially inhibited by heparin.     

Novel anti-inflammatory peptides from the region of highest similarity between uteroglobin and lipocortin I

Significant future developments in the effective treatment of inflammatory diseases may arise from non-toxic dual inhibitors of both cyclooxygenase and lipoxygenase pathways in the arachidonate cascade. Inhibition of phospholipase A2(PLA2)(EC3.1.1.4), may provide such a dual action and recent research has concentrated on the role of PLA2-inhibitory proteins as possible anti-inflammatory agents. Blastokinin or uteroglobin is a steroid-induced rabbit secretory protein with PLA2-inhibitory activity. Its biochemical and biological properties have been extensively studied and its crystallographic structure has been resolved at 1.34 A (refs 15, 16). Lipocortins are a family of related proteins, which, it has been suggested, mediate the anti-inflammatory effects of glucocorticoids (for a review, see ref. 23). Some proteins of this group have been purified and the complementary DNA sequences of two human lipocortins are known. Lipocortins inhibit PLA2 in vitro, although their mechanism of action is still unclear. Recombinant lipocortin I inhibits eicosanoid synthesis in isolated perfused lungs from the guinea pig. Here, we report that synthetic oligopeptides corresponding to a region of high amino-acid sequence similarity between uteroglobin and lipocortin I have potent PLA2 inhibitory activity in vitro and striking anti-inflammatory effects in vivo.     

二(过碘酸)合银(Ⅲ)配离子氧化L-谷氨酰胺的反应动力学及机理

在碱性介质中,二(过碘酸)合银(Ⅲ)配离子([Ag(HIO6)2]5-)氧化Gln导致脱羧与脱氨而生成H2NCOCH2CH2CHO,氧化的产物由化学方法和质谱鉴定.动力学实验表明,氧化反应为总二级,对Ag(Ⅲ)和Gln各为一级,表观二级速率常数随[IO4-]tot增大而减小,随着离子强度的增大而增大,同[OH-]的变化几乎无关.所提出的反应机理包括[Ag(HIO6)2]5-与[Ag(HIO6)(OH)(H2O)]2-形成前期平衡;2种Ag(Ⅲ)的形态被Gln平行还原并且均为速率控制步骤.根据反应机理,导出了速率方程和速控步的速率常数值,计算出了活化参数,对电子转移方式进行了详尽的讨论.     

Protein biosynthesis in organelles requires misaminoacylation of tRNA

In the course of our studies on transfer RNA involvement in chlorophyll biosynthesis, we have determined the structure of chloroplast glutamate tRNA species. Barley chloroplasts contain in addition to a tRNA(Glu) species at least two other glutamate-accepting tRNAs. We now show that the sequences of these tRNAs differ significantly: they are differentially modified forms of tRNA(Gln) (as judged by their UUG anticodon). These mischarged Glu-tRNA(Gln) species can be converted in crude chloroplast extracts to Gln-tRNA(Gln). This reaction requires a specific amidotransferase and glutamine or asparagine as amide donors. Aminoacylation studies show that chloroplasts, plant and animal mitochondria, as well as cyanobacteria, lack any detectable glutaminyl-tRNA synthetase activity. Therefore, the requirement for glutamine in protein synthesis in these cells and organelles is provided by the conversion of glutamate attached to an 'incorrectly' charged tRNA. A similar situation has been described for several species of Gram-positive bacteria. Thus, it appears that the occurrence of this pathway of Gln-tRNA(Gln) formation is widespread among organisms and is a function conserved during evolution. These findings raise questions about the origin of organelles and about the evolution of the mechanisms maintaining accuracy in protein biosynthesis.     

Is sequence conservation in interferons due to selection for functional proteins?

The human alpha-interferon (IFN-alpha) gene family consists of at least 14 potentially functional non-allelic members; the amino acid sequences they encode differ from each other by up to approximately 20% of their residues. Human IFN-beta, which is encoded by a single gene, is distantly related to the IFN-alpha family; it differs in 67% of its residues from IFN-alpha 2. There is considerable evidence that IFN-alpha and -beta compete for the same receptors on their target cells. Comparison of 14 non-allelic human IFN-alpha sequences and the IFN-beta sequence has revealed that 37 of 166 residues are completely conserved and that several of these are arranged in clusters, for example at positions 29-33, 47-50 and 136-150. It is commonly held that evolutionary conservation of amino acids indicates that the residues in question are essential for function. To test this hypothesis in the case of IFNs, we have introduced single site-directed point mutations into the strictly conserved codons 48 and 49 of the IFN-alpha 2 gene which form part of the longest uninterrupted cluster (position 47-50). We report here that the mutant proteins, containing Tyr, Ser and Cys instead of Phe48, or His instead of Gln49, have biological activities indistinguishable from those of wild-type IFN-alpha. In addition, when Glu62, a residue conserved in all known alpha and beta IFNs of man, mouse and cattle, was replaced by Lys, antiviral activity remained unchanged.     

Structure of recombinant human rheumatoid arthritic synovial fluid phospholipase A2 at 2.2 A resolution

Phospholipases A2 (PLA2s) may be grouped into distinct families of proteins that catalyse the hydrolysis of the 2-acyl bond of phospholipids and perform a variety of biological functions. The best characterized are the small (relative molecular mass approximately 14,000) calcium-dependent, secretory enzymes of diverse origin, such as pancreatic and venom PLA2s. The structures and functions of several PLA2s are known. Recently, high-resolution crystal structures of complexes of secretory PLA2s with phosphonate phospholipid analogues have provided information about the detailed stereochemistry of transition-state binding, confirming the proposed catalytic mechanism of esterolysis. By contrast, studies on mammalian nonpancreatic secretory PLA2s (s-PLA2s) have only recently begun; s-PLA2s are scarce in normal cells and tissues but large amounts are found in association with local and systemic inflammatory processes and tissue injury in animals and man. Such s-PLAs have been purified from rabbit and rat inflammatory exudate, from synovial fluid from patients with rheumatoid arthritis and from human platelets. Cloning and sequencing shows that the primary structure of the human s-PLA2 has about 37% homology with that of bovine pancreatic PLA2 and 44% homology with that of Crotalus atrox PLA2. The human s-PLA2 is an unusually basic protein, yet contains most of the highly conserved amino-acid residues and sequences characteristic of the PLA2s sequenced so far. Here we report the refined, three-dimensional crystal structure at 2.2 A resolution of recombinant human rheumatoid arthritic synovial fluid PLA2. This may aid the development of potent and specific inhibitors of this enzyme using structure-based design.     

Gln 对鸭瘟感染鸭十二指肠免疫调节相关基因表达量的影响

摘要: 目的 为谷氨酰胺( Glutamine,Gln) 在正常鸭及鸭瘟( Duck plague,DP) 模型中对肠道功能及作用途径的研究提供依据。方法 280 只无特定病原体( Specific pathogen free,SPF) 7 日龄雏鸭随机分为 8 组,四组梯度剂量每天灌喂 Gln( 0、0. 5、1、2 g / kg 饲 料) 6d,及另四组灌喂同等梯度剂量 Gln 6 d,于 第 一 天 灌 喂 30 min 后 攻 0. 2 mL2000TCID50 鸭瘟病毒,观察 Gln 对正常雏鸭的免疫促进作用及攻毒鸭免疫应激的缓解作用。测定 Toll 样受体 4( Toll-like receptor 4,TLＲ4) 通路基因表达量。结果 Gln 显著提高正常组 TLＲ4 通路 mＲNA 表达量( P ＜ 0. 05) ,显著降低攻毒组表达量( P ＜ 0. 05) 。结论 Gln 通过 TLＲ4 通路提高正常雏鸭肠道的免疫力,降低鸭瘟病毒( Duck plague virus,DPV) 造成的肠道免疫损伤。     

直线式双组分熔体微分电纺“月牙状”聚丙烯纤维制备

搭建直线式熔体微分电纺设备，制备了聚丙烯（PP）和聚乳酸（PLA）双组分纤维，将所制样品放入丙酮水溶液中作浸泡处理，溶解PLA组分获得PP单组分异形纤维；然后对PLA添加增塑剂乙酰柠檬酸三丁酯（ATBC）进行降黏处理，以达到改变PLA熔体流动速率（MFRs）的目的，探究不同MFRs差异所产生的不同包裹现象。研究结果表明，两种组分的MFRs差异对双组分纤维的包裹现象产生显著影响，且通过实验可知，当ATBC质量分数为6%的PLA与PP共纺时，可得到形貌最佳的PP组分异形纤维。     

Crystal structure of a lectin-like natural killer cell receptor bound to its MHC class I ligand

Natural killer (NK) cell function is regulated by NK receptors that interact with MHC class I (MHC-I) molecules on target cells. The murine NK receptor Ly49A inhibits NK cell activity by interacting with H-2D(d) through its C-type-lectin-like NK receptor domain. Here we report the crystal structure of the complex between the Ly49A NK receptor domain and unglycosylated H-2D(d). The Ly49A dimer interacts extensively with two H-2D(d) molecules at distinct sites. At one interface, a single Ly49A subunit contacts one side of the MHC-I peptide-binding platform, presenting an open cavity towards the conserved glycosylation site on the H-2D(d) alpha2 domain. At a second, larger interface, the Ly49A dimer binds in a region overlapping the CD8-binding site. The smaller interface probably represents the interaction between Ly49A on the NK cell and MHC-I on the target cell, whereas the larger one suggests an interaction between Ly49A and MHC-I on the NK cell itself. Both Ly49A binding sites on MHC-I are spatially distinct from that of the T-cell receptor.     

复方北五味子促睡眠口服液镇静催眠作用及机制研究

目的研究复方北五味子促睡眠口服液(Co Schisandra sleep-promoting oral liquid,CSSPOL)的中枢镇静及催眠作用,并初步探讨其作用机制.方法 ICR雄性小鼠40只,随机分为4组,每组10只,分别为空白对照组、CSSPOL低剂量组(4倍稀释液)、中剂量组(2倍稀释液)和高剂量组(原液),按0.1 m L/10 g容量灌胃给药,2次/d,连续7 d.采用小鼠自主活动记录仪观察CSSPOL的中枢镇静作用,采用协同戊巴比妥钠睡眠实验观察其对小鼠的催眠作用,小鼠脑组织中γ-氨基丁酸(γ-aminobutyric acid,GABA)、谷氨酸(glutamate,Glu)和谷氨酰胺(glutamine,Gln)的含量采用酶联免疫吸附法(enzyme-linked immunosorbent assay,ELISA)检测,初步探讨CSSPOL的中枢抑制作用机制.结果 CSSPOL可明显减少小鼠自主活动总次数,缩短阈下剂量戊巴比妥钠诱导小鼠的睡眠潜伏期,并增加睡眠只数,延长阈剂量戊巴比妥钠诱导的小鼠睡眠持续时间,且能使小鼠脑组织中Glu和Gln含量降低.结论 CSSPOL镇静催眠作用显著,其中枢抑制作用可能与降低小鼠脑中兴奋性神经递质Glu和Gln含量有关.     

Phaseolus lunatus Linn vel aff的种子中A型血专一性凝集素的研究

用亲和层析法从Phaseolus lunatus Linn vel aff的种子中纯化出一种对人类A型血专一的凝集素.该凝集素的粗浸提液都只对A型血细胞凝集,而对B、O型绝不凝集.当纯化的凝集素浓度为0.98μg/mL时,即能凝集A型血细胞,GalNAc、对硝基苯酚-β-D-吡喃半乳糖苷能抑制其凝集作用;酚—硫酸法测得凝集素含有3.8％的中性糖;该凝集素是一个促有丝分裂原,其细胞转化为67.9％;它的分子量为56500,有两种亚基,分子量分别为17200和14600.凝集素分子中亮氨酸和缬氨酸含量较高,分别为9.83和9.64,而酸性氨基酸含量远比碱性氨基酸为高.     

Two enzymes bound to one transfer RNA assume alternative conformations for consecutive reactions

In most bacteria and all archaea, glutamyl-tRNA synthetase (GluRS) glutamylates both tRNA(Glu) and tRNA(Gln), and then Glu-tRNA(Gln) is selectively converted to Gln-tRNA(Gln) by a tRNA-dependent amidotransferase. The mechanisms by which the two enzymes recognize their substrate tRNA(s), and how they cooperate with each other in Gln-tRNA(Gln) synthesis, remain to be determined. Here we report the formation of the 'glutamine transamidosome' from the bacterium Thermotoga maritima, consisting of tRNA(Gln), GluRS and the heterotrimeric amidotransferase GatCAB, and its crystal structure at 3.35 A resolution. The anticodon-binding body of GluRS recognizes the common features of tRNA(Gln) and tRNA(Glu), whereas the tail body of GatCAB recognizes the outer corner of the L-shaped tRNA(Gln) in a tRNA(Gln)-specific manner. GluRS is in the productive form, as its catalytic body binds to the amino-acid-acceptor arm of tRNA(Gln). In contrast, GatCAB is in the non-productive form: the catalytic body of GatCAB contacts that of GluRS and is located near the acceptor stem of tRNA(Gln), in an appropriate site to wait for the completion of Glu-tRNA(Gln) formation by GluRS. We identified the hinges between the catalytic and anticodon-binding bodies of GluRS and between the catalytic and tail bodies of GatCAB, which allow both GluRS and GatCAB to adopt the productive and non-productive forms. The catalytic bodies of the two enzymes compete for the acceptor arm of tRNA(Gln) and therefore cannot assume their productive forms simultaneously. The transition from the present glutamylation state, with the productive GluRS and the non-productive GatCAB, to the putative amidation state, with the non-productive GluRS and the productive GatCAB, requires an intermediate state with the two enzymes in their non-productive forms, for steric reasons. The proposed mechanism explains how the transamidosome efficiently performs the two consecutive steps of Gln-tRNA(Gln) formation, with a low risk of releasing the unstable intermediate Glu-tRNA(Gln).     

过度训练对大鼠心肌线粒体MDA，SOD，GSH—Px，PLA2及Ca^2＋浓度的影响

为探讨过度训练状态下心肌组织损伤的变化规律,笔者采用过度训练大鼠模型,将40只大鼠随机分为训练对照组和过度训练组,分别进行8周的中等强度和大强度跑台训练,于第8周周末运动后处死全部大鼠,取心室肌测定线粒体丙二醛、超氧化物歧化酶、谷胱苷肽过氧化物酶、磷脂酶A：及Ca^2＋浓度．结果表明：过度训练状态下,大鼠心肌线粒体丙二醛含量、磷脂酶A2活性及Ca^2＋浓度显著升高,超氧化物歧化酶、谷胱苷肽过氧化物酶活性显著下降．过度训练后,大鼠心肌线粒体自由基产生增加,抗氧化能力下降,引起线粒体膜脂质过氧化水平增加,膜降解反应增强,从而导致大鼠心肌线粒体损伤和心肌组织的损伤．     

Platelet activation--a role for a 40K anti-phospholipase A2 protein indistinguishable from lipocortin

Stimulus-response (S-R) coupling in platelets requires an intermediary other than an elevation in cytosolic free calcium ([Ca2+]i). While an increase in [Ca2+]i is essential in S-R coupling, effecting phosphorylation of myosin of relative molecular mass (Mr) 20,000 (20 K), platelet activation is also associated with phosphorylation of a 40K protein, which can occur in the absence of changes in [Ca2+]i. The 40K protein is the substrate for protein kinase C (PKC). Mounting evidence suggests that activation of PKC by diacylglycerol is the other signal involved in S-R coupling. Although phosphorylation of the 40K protein is associated with certain platelet functional responses, no precise role has been accredited to it. Recently, we and others have described several proteins (collectively known as lipocortin) which inhibit phospholipase A2 (PLA2). One of the most conspicuous proteins of this group is a 40K peptide whose inhibitory activity can be suppressed by prior phosphorylation. We hypothesized that the 40K protein described in platelets may possess anti-PLA2 activity and that phosphorylation by PKC, suppressing its inhibitory activity, may represent the mechanism underlying mobilization of arachidonic acid, the precursor of prostaglandins. The results of the present study strongly support this hypothesis.     

补充谷氨酰胺对大学生暑期集训期血清T/C值、血尿素的影响

探讨了补充谷氨酰胺对暑期大运动量训练的大学生运动员血清T/C值、血尿素变化的影响,旨在进一步揭示谷氨酰胺调节机体蛋白质代谢的作用机制。结果显示:谷氨酰胺能明显抑制男生运动性低血睾发生,使T/C值维持在相对高的水平,男生组间T/C值出现了显著性差异(P?0.05),本实验中谷氨酰胺对女生血睾酮及T/C值变化作用不明显;谷氨酰胺对大学生实验期间血尿素变化没有显著影响。提示:适量补充谷氨酰胺能够通过影响T/C值,有利于机体蛋白质的合成代谢,对于保持人体在长时间、大运动量训练时较好的机能状态,延缓疲劳发生有积极的生物学作用。     

纳米HMCM-49分子筛上1-己烯异构化/芳构化

采用低温老化,高温晶化两步法在Na₂O-Al₂O₃-SiO₂-HMI-H₂O体系中水热动态合成了纳米MCM-49分子筛,采用XRD,SEM,N₂吸/脱附曲线和NH₃-TPD等手段对MCM-49分子筛结构和酸性质进行了表征,并在固定床微反应器上评价了HMCM-49分子筛对1-己烯异构化和芳构化催化性能。结果表明与微米HMCM-49分子筛相比,纳米HMCM-49分子筛比表面积明显增大,强酸量和总酸量有所下降。纳米分子筛具有更佳的异构化催化活性,异构化及芳构化产物总收率高于微米分子筛且积碳量少。     

聚乳酸的酶降解动力学模型

将连续分布理论应用到聚乳酸的酶降解反应动力学中,模拟蛋白酶K降解聚乳酸的过程。将聚乳酸的酶降解过程分为3步:酶吸附在聚乳酸上;聚乳酸和酶形成具有活性的中间过渡体;生成特定断裂产物。运用矩的运算将积分微分方程转变成便于计算的普通微分方程,计算得到的降解产物的质量浓度随时间的变化与实验基本吻合。模型参数(k′,kE)与酶浓度、流量、pH和温度变化有关。     

聚乳酸微球制备的初步研究

聚乳酸PLA(polylactide)是一种无毒、可生物降解的聚合物,它具有良好的生物相容性,在医药上有广泛的应用.通过实验研究了聚乳酸浓度,表面活性剂浓度以及两者的配料比对溶媒挥发法制备聚乳酸微球及微球粒经的影响.实验为进一步制备医用聚乳酸微球和类似的医用药剂做了有益的探索.     

Preparation and cytocompatibility of polylactic acid/hydroxyapatite/graphene oxide nanocomposite fibrous membrane

A series of polylactic acid (PLA) based nanocomposite fibrous membranes,including neat PLA,PLA/hydroxyapatite (HA) and PLA/HA/graphene oxide (GO),were fabricated via electrospinning method.The morphology and composition were investigated by scanning electron microscopy (SEM),X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) respectively.The thermal stability was determined by thermogravimetric analysis (TGA).To estimate the cytocompatibility of asprepared PLA/HA/GO fibrous membrane,MC3T3-E1 cells were cultured,and the corresponding cell adhesion and differentiation capability were investigated by fluorescence microscopy,SEM and MTT test.The electrospun ternary PLA/HA/GO membrane exhibited three-dimensional fibrous structure with relatively rough surface morphology,which made itself ideal for cell attachment and proliferation in bone tissue regeneration.The fluorescence microscopy,SEM and MTT test confirmed that the PLA/HA/GO nanocomposite fibrous membrane created a proper environment for the seeding and proliferation of MC3T3-E1 cells.