IBV青岛腺胃分离株（SD／97／02）S1蛋白基因的序列测定和分析

参考Genbank上发表的IBV S1纤突蛋白基因序列,设计了一对引物,对鸡传染性支气管炎病毒青岛腺胃分离株（SD／97／02）RNA进行RT－PCR扩增。将PCR产物克隆入pMD18-T载体中进行序列测定和分析。序列分析表明,该毒株的S1基因的G＋C％含量较少,为37.0%,存在HindⅢ,BamHⅠ,BglIⅠ,SacⅠ和SalⅠ位点,无EcoRⅠ位点,与其他毒株的同源性在87.02%－94.21%之间,在第154－429nt处为高度的变异区;将基因序列翻译成氨基酸后,假定的S1蛋白由540个氨基酸组成,等电点8.24,在蛋白质内部存在18个Cys,在S1与S2蛋白之间的剪切位点为HRRRR,这与大多数IBV毒株（RRF／SRR）不一样,有三个区域的氨基酸序列高度保守;169－181aa,230-250aa,485-506aa;与其他毒株进行抗原性比较后发现,在该毒株的320－326aa及390－401aa处的抗原表位消失,而在325－345aa、379－389aa处则出现了很强的抗原表位;第438－444aa处,其他IBV毒株（除ZJ971株外）原来存在的强抗原位点在本毒株中消失。在53－65位的氨基酸抗原性与其他毒株相比明显变弱。     

鸡传染性支气管炎病毒中国分离株LX4纤突蛋白基因分子特征

应用RT-PCR方法分别扩增了中国地方分离株IBV LX4 S1和S2基因并进行了基因的克隆和序列测定.结果发现,LX4 S基因由3495个核苷酸组成,编码一条1164个氨基酸残基组成的多肽.S基因编码产物裂解后形成的S1和S2亚单位分别由539和625个氨基酸残基组成.LX4 S基因推导氨基酸切割识别位点序列为HRRRR,与A2、SD/97/02和Z株相同,而与其它国内外参考毒株不同.与国内外10株已报道的具有全S基因序列的IBV参考毒株比较,LX4与国内分离毒株SD/97/02的核苷酸和氨基酸同源性最高.与32株国内外参考毒株的S1基因进化树分析比较表明,LX4与A2、SD/97/02、Z、TJ/96/02、JX/99/01和SAIBWJ等7个国内分离株在同一亚群内.在该亚群内,LX4与A2和SD/97/02亲缘关系更近,且三者在高变区和抗原表位均具有高度的同源性,而与本亚群内其它参考毒株对应的高变区和抗原表位同源性差异较大.LX4与H120 S1基因编码氨基酸的同源性虽然高于其它国外参考毒株,但同源性仍然较低,为75%.与参考毒株比较,LX4 S2基因的点突变造成其推导的氨基酸序列有11个位点发生改变,这些突变可能影响S2与S1蛋白之间的相互作用,从而影响S蛋白与特异性抗体的结合.     

鸡传染性支气管炎病毒两个地方分离株S1基因的扩增克隆与序列分析

对IBV肾型毒株JS/95 /0 3和呼吸型毒株SD/97/0 1的S1全基因进行了扩增、克隆和序列测定 ,将两毒株的S1基因序列与 10个参考毒株进行了比较。结果表明 ,JS/95 /0 3和SD/97/0 1分别与M41和H12 0株亲缘关系最近 ,它们可能是疫苗毒株的变异株。JS/95 /0 3和SD/97/0 1间的亲缘关系也较近 ,两者S1蛋白中只有 2 4个氨基酸的差异 ,其中有 15个位于前 130个氨基酸中 ,第 116位氨基酸可能与毒株的致病性有关。对两毒株S1蛋白的二级结构进行了预测和比较 ,结果发现 ,一个或极少数氨基酸的差异即可导致S1蛋白二级结构和抗原性的改变     

鸡传染性支气管炎病毒两个地方分离株S1基因的扩增克隆与序列分析

对IBV肾型毒株JS/95/03和呼吸型毒株SD/97/01的S1全基因进行了扩增、克隆和序列测定,将两毒株的S1基因序列与10个参考毒株进行了比较.结果表明,JS/95/03和SD/97/01分别与M41和H120株亲缘关系最近,它们可能是疫苗毒株的变异株.JS/95/03和SD/97/01间的亲缘关系也较近,两者S1蛋白中只有24个氨基酸的差异,其中有15个位于前130个氨基酸中,第116位氨基酸可能与毒株的致病性有关.对两毒株S1蛋白的二级结构进行了预测和比较,结果发现,一个或极少数氨基酸的差异即可导致S1蛋白二级结构和抗原性的改变.     

中国1995～2004年鸡传染性支气管炎病毒地方分离株膜蛋白基因的遗传变异分析

应用RT-PCR方法扩增到了我国1995～2004年20株IBV现地分离株的膜蛋白(Membrane,M)基因片段.序列测定表明,20株IBV分离株M基因开放阅读框由672～681bp组成,编码由223～226个氨基酸残基组成的多肽.与我国分离株LX4相比,M基因推导氨基酸序列的变异主要发生在2～17位、221～223位,其中4～6位存在氨基酸的插入和缺失,导致IBV毒株间M蛋白糖基化位点的差异.与GenBank中34株IBV参考毒株M蛋白基因推导氨基酸序列进行比较和分析,系统进化关系显示54株IBV毒株分属于5个进化群.我国IBV分离株M基因在进化关系上较为独立,主要分布在第Ⅱ群和第Ⅳ群,其中第Ⅱ群分离株和中国台湾毒株进化关系密切.此外,参考IBV国内分离株S1基因及N基因系统发育进化树的研究结果,并与M基因进行比较,表明我国IBV也存在着基因重组现象,尤其是疫苗毒和流行毒之间的重组.     

猪流行性腹泻病毒CH/JL毒株S基因的克隆、序列分析及线性抗原表位区的鉴定

以猪流行性腹泻病毒CH/JL毒株的RNA为模板,通过RT-PCR扩增获得的3个相互重叠的cDNA克隆覆盖了S基因,序列比对结果表明:PEDV CH/JL株S基因与CV777、Brl/87、JS、KPEDV和Chinju99毒株S基因核苷酸序列的同源性分别为96.97%、96.87%、96.41%、94.02%和93.93%,氨基酸序列的同源性分别为96.17%、95.88%、96.10%、92.36%和92.05%;分子进化树分析结果显示,PEDV CH/JL株S基因与JS毒株S基因亲缘关系最近,处于同一群。利用DNAstar Protean程序预测了PEDV CH/JL株S蛋白一个抗原表位区(83~276aa),将其克隆到原核表达载体pGEX-6p-1后转化E.coliBL21(DE3)感受态细胞,在终浓度1.0mmol/L的IPTG诱导下获得了表达,Western blot结果显示,预测的抗原表位区GST融合蛋白能与猪流行性腹泻病毒多克隆抗血清反应,提示该抗原表位区含有线性抗原表位。     

中国西藏小反刍兽疫病毒野生株China/Tib/Gej/07-30基质蛋白基因和融合蛋白基因的分子特征分析

对我国西藏小反刍兽疫病毒野生株China/Tib/Gej/07-30进行基质蛋白(M)和融合蛋白(F)基因序列测定,并进行分子生物学特征分析。首先应用逆转录聚合酶链式反应扩增出M和F基因片段,对聚合酶链式反应产物进行直接测序,然后对测定的核苷酸和推测的氨基酸序列进行比较分析。China/Tib/Gej/07-30的M基因由1483个核苷酸组成,编码335个氨基酸,与其他分离株核苷酸和氨基酸序列同源性分别为92.4%～97.7%和97.0%～98.2%。F基因由2411个核苷酸组成,编码546个氨基酸,与其他分离株核苷酸和氨基酸序列同源性分别为85.5%～96.1%和94.3%～98.2%。China/Tib/Gej/07-30的F蛋白含有信号肽序列和跨膜结构域,序列高度变异。F蛋白第104～108位和第109～133位氨基酸位点分别是高度保守的裂解位点和融合肽结构域。F蛋白还含有序列高度保守的三个七肽重复区。China/Tib/Gej/07-30的M基因3′端的非编码区(UTR)长度为443个核苷酸,GC含量高达68.4%,与其他PPRV毒株的同源性为82.4%～93.5%。China/Tib/Gej/07-30的F基因5′UTR区长度为634个核苷酸,GC含量高达70.0%,与其他PPRV毒株序列相似性为76.2%～91.7%。     

犬冠状病毒大熊猫株纤突蛋白全基因的克隆与序列分析

在国内首次对犬冠状病毒大熊猫野毒株(CCV DXMV)纤突蛋白基因进行了克隆和序列测定.该基因全长4362bp,编码1453个氨基酸,N端前18个氨基酸为推测的信号肽序列,后1435个氨基酸构成成熟蛋白.与GenBank中已发表的11个CCV毒株S基因相比,S基因核苷酸序列同源性在40.2%～99.5%之间;推导的氨基酸序列同源性在15.9%～99.0%之间.DXMV株S基因变异区主要集中在该基因前1/2处,其中350-370、439-478、1718-1818三个区域碱基变异较大,而1060-1700区却十分保守.基于S全基因及其蛋白的聚类分析表明,DXMV株与K378、NVSL和US patent株亲源关系最近.推导的DXMV株S蛋白氨基酸序列潜在的N-联糖基化位点与CCV强毒V54相同,为34个,比Insavc-1弱毒多一个;其中第566-568位糖基化位点为多数强毒拥有而弱毒没有的.另外,DXMV株S蛋白疏水性及抗原表位与其它毒株有一定的差异,这些差异对DXMV株致病性和免疫原性等影响尚待进一步的研究.     

表达轮状病毒SA11株Vp4的抗原表位诱导病毒中和抗体生成

以昆虫病毒Flockhousevirus（FHV）外壳蛋白为载体的外源抗原表位表达系统（FHV-RNA2载体系统）．在重组杆状病毒和重组pET系统中构建和表达了SA11Vp4胰酶切割位点两侧和重叠切割位点3个抗原表位氨基酸序列（抗原表位A,aa223~242;抗原表位B,aa243～262;抗原表位C,aa234～251）,并对其免疫原性进行了研究。结果表明：这3个抗原表位能诱导动物产生抗同源氨基酸序列的抗体和抗同源病毒（SA11）感染性的血清中和抗体。研究结果提示：RVVp4胰酶切割位点区氨基酸序列除了具有胰酶切割增强病毒感染力外,还具有诱导动物机体产生血清中和抗体的能力,是RV重组抗原表位亚单位疫苗研究中重要的抗原表位氨基酸序列。     

广东地区流感H3N2毒株血凝素基因特征、进化和变异分析

为揭示广东地区2007～2010年甲型H3N2毒株血凝素(HA)基因特征和变异,采用时空抽样方法抽样,检测广东2007～2010年甲型H3N2毒株HA基因核苷酸序列,同时检索全球HA基因序列作为对照,采用Lasergene 7.1和Mega 5.05软件对HA基因核苷酸序列进行比对和分析;并结合流行病学资料,对变异毒株进行进化速度分析;同时进行抗原分析。结果发现,广东2007～2010年H3N2毒株HA基因同义进化(Ks)和错义进化(Ka)速度分别为2.06×1E-3～2.23×1E-3核苷酸/年和1.05×1E-3～1.21×1E-3核苷酸/年,HA1较HA2的错义突变速率要高3.13倍。与疫苗株A/Perth/16/2009的HA基因比较,2009年广东毒株同源性达到98.8%～99.7%、2010年同源性达到98.0%～98.4%。在广东2007～2010年毒株中,HA1五个抗原表位均有氨基酸位点变异,尤其是2010年毒株B区(N160K)和D区(K174R/N)的变异;此外,广东2010年毒株受体结合部位(RBS)还发生K189E/N/Q和T228A置换变异;两个糖基化位点变异影响到抗原性;目前使用的H3N2疫苗株与目前流行毒株的抗原性有差异。广东地区2007～2010年的毒株中,血凝抑制抗体的抗原分析结果有差异。结果提示,目前广东乃至全球甲型H3N2毒株HA1B区和D区均有氨基酸位点变异,RBS的两个位点发生置换,糖基化位点变异影响到表位A区和B区抗原性;与WHO推荐2011年流感H3N2毒株疫苗株比较,目前流行毒株HA基因有抗原位点变异。     

ATP/ADP exchange activity of gastric (H+ + K+)-ATPase

The ATP/ADP exchange is shown to be a partial reaction of the $\text{(}\text{H}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ by the absence of measurable nucleoside diphosphokinase activity and the insensitivity of the reaction to $\text{P}{}^1$, $\text{P}{}^5$ -di(adenosine-5′) pentaphosphate, a myokinase inhibitor. The exchange demonstrates an absolute requirement for Mg²⁺ and is optimal at an ADP/ATP ratio of 2. The high ATP concentration ($\text{K}{}_{0.5}\text{= 116 μ}\text{M}$) required for maximal exchange is interpreted as evidence for the involvement of a low affinity form of nucleotide site. The ATP/ADP exchange is regarded as evidence for an ADP-sensitive form of the phosphoenzyme. In native enzyme, pre-steady state kinetics show that the formation of the phosphoenzyme is partially sensitive to ADP while modification of the enzyme by pretreatment with 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) in the absence of Mg²⁺ results in a steady-state phosphoenzyme population, a component of which is ADP sensitive. The ATP/ADP exchange reaction can be either stimulated or inhibited by the presence of K⁺ as a function of pH and Mg²⁺.     

A/California/7/2009与A/California/4/2009病毒感染力比较

目的甲型H1N1流感病毒A/California/7/2009与A/California/4/2009病毒序列比较同源性在99%以上,本实验旨在比较两株病毒感染BALB/c小鼠研究感染力强弱。方法分别将A/California/7/2009（CA7）与A/California/4/2009（CA4）两株病毒分别连续10倍稀释后,对4~6周龄雌性BALB/c小鼠经乙醚麻醉后进行滴鼻攻毒,每个稀释度接种10只实验小鼠,测定CA7 MLD50为101.24/0.05 mL,检测小鼠感染、致病的多项指标,观察期为14 d。结果相同TCID50的CA7和CA4病毒感染小鼠,CA4感染小鼠后14 d内死亡率为20%,而CA7感染小鼠后8 d内死亡率为100%。CA7 106TCID50感染的小鼠病理表现为重度弥漫性间质性肺炎,CA4 106TCID50感染的小鼠病理表现为中度-重度间质性肺炎。结论在相同条件下,CA7感染力明显强于CA4。     

AutoFACT: AnAutomaticFunctionalAnnotation andClassificationTool

Background  

Assignment of function to new molecular sequence data is an essential step in genomics projects. The usual process involves similarity searches of a given sequence against one or more databases, an arduous process for large datasets.     

Comparison of parameters estimated from A/Ci and A/Cc curve analysis

The parameters estimated from traditional A/C _i curve analysis are dependent upon some underlying assumptions that substomatal CO₂ concentration (C _i) equals the chloroplast CO₂ concentration (C _c) and the C _i value at which the A/C _i curve switches between Rubisco- and electron transport-limited portions of the curve (C _i-t) is set to a constant. However, the assumptions reduced the accuracy of parameter estimation significantly without taking the influence of C _i-t value and mesophyll conductance (g _m) on parameters into account. Based on the analysis of Larix gmelinii’s A/C _i curves, it showed the C _i-t value varied significantly, ranging from 24 Pa to 72 Pa and averaging 38 Pa. t-test demonstrated there were significant differences in parameters respectively estimated from A/C _i and A/C _c curve analysis (p<0.01). Compared with the maximum ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) carboxylation rate (V_cmax), the maximum electron transport rate (J_max) and J_max/V_cmax estimated from A/C _c curve analysis which considers the effects of g _m limit and simultaneously fits parameters with the whole A/C _c curve, mean V_cmax estimated from A/C _i curve analysis (V_cmax-C _i) was underestimated by 37.49%; mean Jmax estimated from A/C _i curve analysis (J_max-C _i) was overestimated by 17.8% and (J_max-C _i)/(V_cmax-C _i) was overestimated by 24.2%. However, there was a significant linear relationship between V_cmax estimated from A/C _i curve analysis and V_cmax estimated from A/C _c curve analysis, so was it J_max (p<0.05).     

Characterization of cytochrome P-450SCC-containing liposomes

Purified cytochrome $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ from bovine adrenocortical mitochondria was incorporated into liposomes by the cholate-dilution method utilizing either dialysis or Sephadex gel filtration. Among synthetic phospholipids tested, dioleoylglycerophosphocholine showed the best stability during the incorporation of $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ into liposomes. A maximum amount of heme was incorporated into liposomes at a molar ratio of phospholipid to the cytochrome of approx. 200. When $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ was incorporated into the dioleoylglycerophosphocholine liposomes by the cholate-filtration method, the $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}\text{-}\text{containing liposomes}$ showed two major populations on the elution pattern of the Sepharose 4B gel filtration, and were seen at a diameter of 200–600 Å and its aggregated forms. When the cytochrome was incorporated into dioleoylglycerophosphocholine liposomes or cholesterol-free adrenocortical mitochondrial liposomes, $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ was less stable than $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ in aqueous solution. Cholesterol or adrenodoxin markedly stabilized the liposomal $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$. Liposomal $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ required cholesterol for its optimum reduction with adrenodoxin, adrenodoxin reductase, and NADPH in the presence of CO. About 70% of the total heme in the dioleoylglycerophosphocholine liposomes was reduced by the enzymatic reduction in the presence of cholesterol, indicating that 70% of the total molecules are exposed to the surface of the outer monolayer. In order to see the location of the heme in membrane, the dioleoylglycerophosphocholine-liposomal $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$ was subjected to $\text{p-}\text{chloromercuriphenyl sulfonic acid}$ treatment. This reagent destroyed the liposomal $\text{P}{}_{\mathrm{<mtext>450</mtext><mtext>SCC</mtext>}}$. These results suggest that the heme is located in the proximity of the $\text{p-}\text{chloromercuriphenyl sulfonic acid}$ reacting sites which are exposed to the surface, or located on the vincinity of polar heads of the membrane.     

Ligand-dependent reactivity of (Na+ + K+)-ATPase with showdomycin

Showdomycin inhibited pig brain ($\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ with pseudo first-order kinetics. The rate of inhibition by showdomycin was examined in the presence of 16 combinations of four ligands, i.e., Na⁺, K⁺, Mg²⁺ and ATP, and was found to depend on the ligands added. Combinations of ligands were divided into five groups in terms of the magnitude of the rate constant; in the order of decreasing rate constants these were: (1)$\text{Na}{}^{\mathrm{+}}\text{+}\text{Mg}{}^{\mathrm{2+}}\text{+}\text{ATP}$, (2) $\text{Mg}{}^{\mathrm{2+}}$, $\text{Mg}{}^{\mathrm{2+}}\text{+}\text{K}{}^{\mathrm{+}}$, $\text{K}{}^{\mathrm{+}}$ and none, (3) $\text{Na}{}^{\mathrm{+}}\text{+}\text{Mg}{}^{\mathrm{2+}}\text{,}\text{Na}{}^{\mathrm{+}}$, $\text{K}{}^{\mathrm{+}}\text{+}\text{Na}{}^{\mathrm{+}}$ and $\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{+}\text{Mg}{}^{\mathrm{2+}}$, (4) $\text{Mg}{}^{\mathrm{2+}}\text{+}\text{K}{}^{\mathrm{+}}\text{+}\text{ATP}\text{,}\text{K}{}^{\mathrm{+}}\text{+}\text{ATP}$ and $\text{Mg}{}^{\mathrm{2+}}\text{+}\text{ATP}\text{, (5)}\text{K}{}^{\mathrm{+}}\text{+}\text{Na}{}^{\mathrm{+}}\text{+}\text{ATP}$, $\text{Na}{}^{\mathrm{+}}\text{+}\text{ATP}$, $\text{Na}{}^{\mathrm{+}}\text{+}\text{ATP}$, $\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{+}\text{Mg}{}^{\mathrm{2+}}\text{+}\text{ATP}$ and ATP. The highest rate was obtained in the presence of Na⁺, Mg²⁺ and ATP. The apparent concentrations of Na⁺, Mg²⁺ and ATP for half-maximum stimulation of inhibition ($\text{K}{}_{0.5}{}^{\mathrm{<mtext>s</mtext>}}$) were 3 mM, 0.13 mM and 4μM, respectively. The rate was unchanged upon further increase in Na⁺ concentration from 140 to 1000 mM. The rates of inhibition could be explained on the basis of the enzyme forms present, including E₁, E₂, ES, E₁-P and E₂-P, i.e., E₂ has higher reactivity with showdomycin than E₁, while E₂-P has almost the same reactivity as E₁-P. We conclude that the reaction of ($\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ proceeds via at least four kinds of enzyme form (E₁, E₂, E₁ · nucleotide and EP), which all have different conformations.     

Characterization of partially purified (Na+ + K+)-ATPase from porcine lens

The partial purification of $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ from pig lens has been achieved by treatment with deoxycholate followed by density gradient centrifugation. The specific activity of the final preparation, ranging from 300 to 500 nmol/h per mg protein, is increased approx. 100-fold compared to the homogenate. A parallel increase in $\text{p-}\text{nitrophenylphosphatase}$ activity is also observed. Sodium dodecyl sulfate (SDS) gel electrophoresis reveals six major protein bands, one of which is the 93 kDa α subunit of $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ which can be phosphorylated by reaction with [γ-³²P]ATP. A second band contains a glycoprotein which displays an apparent molecular weight of 51 000 and thus appears to be the β subunit of the enzyme. The enzyme is sensitive to ouabain with the $\text{I}{}_{50}$ for $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ and $\text{p-}\text{nitrophenylphosphatase}$ inhibition being 1.2 and 1.3 μM, respectively. Several agents which inhibit $\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ from other tissues such as oligomycin, Ca²⁺, vanadate, $\text{N-}\text{ethylmaleimide}$, $\text{p-}\text{chloromercuribenzenesulfonic acid}$ (PCMBS) and 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB) also inhibit the lens enzyme. Monovalent cations other than K⁺ are partially effective in activating the ($\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}$)-ATPase and $\text{p-}\text{nitrophenylphosphatase}$ activities. The K⁺ congeners were relatively more effective in supporting $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ compared to $\text{p-}\text{nitrophenylphosphatase}$ activity. Other kinetic properties of the lens enzyme are also comparable to those of the enzyme from other tissues. Utilizing the partially purified membrane bound enzyme, discontinuities in Arrhenius plots of $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ activity, $\text{p-}\text{nitrophenylphosphatase}$ activity and fluoresence polarization of the fluidity probe, 1,6-diphenyl-1,3,5-hexatriene (DPH), are observed near the physiological temperature of lens. The possible significance of these observations for the mechanism of cataract formation are discussed.     

An electron spin probe study of (Na+ + K+)-ATPase-Containing membranes

The modulating effect of membrane lipids on enzyme function has been described by several investigators. We have used the spin probe $\text{N-}\text{oxyl}\text{-4′,4′-}\text{dimethyloxazolidine}\text{-12-}\text{keto}$ methyl stearate (M 12-NSE) to study this interaction in ox brain membranes enriched with $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$. This methyl ester of stearic acid is practically insoluble in aqueous media, and consequently spectra of M 12-NSE-labelled preparations are free of “liquid lines”.At least two types of spectra may be obtained when ox brain microsomes are spin labelled with M 12-NSE, indicating the presence of two distinct binding sites. At one site the spin label is relatively unrestricted and gives rise to an isotropic spectrum. A second spectrum, which is obtained from spin label at another site, is similar to that which is observed after incorporation of M 12-NSE into phospholipid bilayers. This suggests that this latter site is within the core of the microsomal membrane.The two binding sites differ in their affinity for the spin probe. The low affinity site is both more abundant in crude preparations and is more easily removed by detergent treatment; spin labels at this site produce isotropic spectra. The high affinity sites are fewer in number and produce broad spectra. In addition these high affinity sites increase in concentration as the enzyme undergoes purification.The two sites are quite distinct in their sensitivity to ascorbic acid, the low affinity site showing a considerably greater rate of reduction by this agent.This study also demonstrates that the delipidation effects of sodium dodecyl sulfate and sodium deoxycholate on $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase-enriched}$ microsomes from ox brain are not identical.It is suggested that the two spin probe binding sites represent two different lipid domains, one of which is very closely associated with the $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ enzyme and may reflect a protein-directed phospholipid specificity for this enzyme.     

Occurrence of N-malonyl-D-alanine in pea seedlings

One of the ninhydrin-negative alanine conjugates isolated from pea seedlings was identified as $\text{N-}\text{malonyl}\text{-}\text{D}\text{-}\text{alanine}$.The identification of this conjugate was carried out by a comparison of its gas-liquid chromatographic and mass spectrometric properties, and its nuclear magnetic resonance and infrared spectra with those of synthetic $\text{N-}\text{malonyl}\text{-}\text{D}\text{-}\text{alanine}$. The alanine in the conjugate was shown to be present as the D-isomer by enzymatic and chromatographic analyses.     

catmap: Case-control And TDT Meta-Analysis Package

Background  

Risk for complex disease is thought to be controlled by multiple genetic risk factors, each with small individual effects. Meta-analyses of several independent studies may be helpful to increase the ability to detect association when effect sizes are modest. Although many software options are available for meta-analysis of genetic case-control data, no currently available software implements the method described by Kazeem and Farrall (2005), which combines data from independent family-based and case-control studies.