表面活性剂的疏水性及电性对肌酸激酶的活力及复性能力的影响

测定了烷基硫酸钠(CnH_(2n+1)SO_4Na;记为CnS;n=8,10,12)和溴化十烷基三甲铵C_(10)H_(21)(CH_3)Br;记为C_(10)NM_3)对肌酸激酶(CreatineKinase;记为C.K.)的活力,以及在它们中变性后复活能力的影响。CnS对C.K.的变性效率随n的增加而增加,变性效率的对数和n之间有线性关系;CnS水C.K.的变性能力远大于C_(10)NM_3;C.K.被C_(10)NM_3变性以后,其复性能力(稀释时恢复活力的程度远大于C.K.被CnS变性后活力的恢复能力。这种差别主要是由于C_(10)NM_3带正电,而CnS带负电引起的。     

溴化十烷基三甲铵对肌酸激酶的变性及复性

测定了溴化十烷基三甲铵(C_(10)H_(21)N(CH_3)_3Br;记为C_(10)NM_3)对肌酸激酶(Creatine Kinase;记为C.K.)的活力及构象的影响,以及变性后C.K.的复性。实验结果表明:C_(10)NM_3对C.K.有很强的变性能力,在0.06M时,就可以使C.K.完全失活;在0.08M时,就可使C.K.内部的6个巯基有4个暴露出来;与SDS变性剂不同,C.K.在高浓度的C_(10)NM_3中变性以后,直接冲稀时就可以完全复性。FTIR以及CD等实验方法证明,尽管C_(10)NM_3能使C.K.的构象发生明显变化,但C.K.的二级结构几乎不受C_(10)NM_3的影响。     

盐酸己烷基双胍对肌酸激酶的活力及构象的影响

测定了盐酸己烷基双胍(C_6H_(13)C_2N_5H_(?)HCl;记为HBGC)对肌酸激酶(Creatine Kinase;记为CK)的活力的影响;同时利用荧光光谱,紫外差光谱.付里叶红外光谱等手段测定了HBGC对CK构象的影响,结果表明.HBGC为0.07mol时就使CK完全失活、而且在高浓度的HBGC中变性后的CK.稀释时即可完全复活;CK的活力丧失明显先于构象的变化,与溴化+烷基三甲铵不同,HBGC可以破坏CK的二级结构,使之完全无序化.     

底物对变性酶的修复作用

 兎肌肌酸激酶被LDS变性后,底物能够诱导变性酶使其活力和构象得到部分恢复。变性程度不同的酶,构象和活力的恢复程度也不同:低浓度LDS变性酶,恢复程度较高;反之亦然。活力的恢复与构象的恢复两者呈对应关系。底物修复作用的pH以8.2为好。底物修复作用受其它蛋白质(例如BSA)存在的影响。等速电泳结果表明,BSA能竞争性结合LDS-酶复合物的LDS,使酶成为游离酶。变性酶先与BSA保温再加底物所得的活力恢复,大约是变性酶与含BSA的底物保温所得活力的10倍。这一结果似表明LDS变性酶仍能结合底物;被结合的底物还能使变性酶的构象发生变化。     

烷基聚氧乙烯醚硫酸钠对肌酸激酶的变性及复性

系统的研究了十二烷基聚氧乙烯醚硫酸钠Ｃ１２Ｈ２５（ＯＣ２Ｈ４）ｎＳＯ４Ｎａ（ｎ＝０,１,３,５,７;记为Ｃ１２ＥｎＳ）对肌酸激酶的活力和构象的影响。结果表明：随着氧乙烯基个数（ｎ）的增加,Ｃ１２ＥｎＳ对Ｃ．Ｋ．的活力破坏逐步减小;Ｃ１２ＥｎＳ引起Ｃ．Ｋ活力的丧失明显早于可测构象的变化。ＣＫ经Ｃ１２ＥｎＳ变性后,稀释时即可完全复性。Ｃ１２Ｅ７Ｓ不仅对Ｃ．Ｋ的变性能力强（１０ｍＭ就使Ｃ．Ｋ完全失活）;而且在１０－９５℃的温度范围内,能阻止Ｃ．Ｋ聚集沉淀。Ｃ１２ＥｎＳ是Ｃ．Ｋ的高效可逆变性剂。     

底物对LDS变性肌酸激酶的修复作用

本文报导了底物能够诱导变性肌酸激酶活力和构象部分恢复的实验事实.92μm免肌酸激酶用47mMLDS在pH9.0的甘氨酸缓冲液中变性半小时,取1μl变性酶至2ml底物系统中,10分钟后可观察到酶活力的再现:半小时可达对照酶活力的10%.当把此变性酶液按同样倍数稀释至与测活系统相同的缓冲液中,不同时间取样测活,则观察不到活力再现.可见,前述活力再现来自底物对变性酶的作用.底物对变性酶修复后的活力随着变性程度的减小而增大.底物可使低浓度LDS变性酶活力恢复至天然酶水平.ATP对构象修复起着主要作用.     

按摩对兔骨骼肌损伤修复自由基相关酶的活性的影响

目的通过检测按摩对兔骨骼肌钝挫伤修复过程中谷胱甘肽过氧化物酶(GSH-Px)、髓过氧化物酶(MPO)活力值,及细胞磷脂酶A2(PLA2)mRNA表达的影响,探讨按摩加快清除肌肉损伤修复过程中自由基的作用机制。方法选取新西兰大白兔42只,随机分为4组,即正常对照组(A组,n=3)、损伤组(B组,n=3)、按摩组(C组,n=18,包括损伤后第5d和第10d各9只)和自然恢复组(D组,n=18,包括损伤后第5d和第10d各9只)。A组实验兔不作处理,作为正常对照;B、C、D组实验动物用自制打击器来制备兔左后肢股四头肌损伤模型。C组于造模后第4天用按摩器施以按摩治疗,1次/天至处死取材;B、D组不进行按摩。各组于伤后5d及10d活体心脏取血,用于进行生化指标检测GSH-Px和MPO的活力值;取股四头肌样本,进行HE染色和实时荧光定量PCR法检测PLA2mRNA的相对表达量,并同A组正常股四头肌检测结果进行比较。结果正常组、损伤组、伤后5d及10d按摩组和自然恢复组的GSH-Px和MPO活力值经比较,按摩组较自然恢复组明显减少(P0.05);经按摩干预后,各组的PLA2mRNA表达量也显示,按摩组明显比自然恢复组减少(P0.05)。结论按摩可促进兔股四头肌损伤的修复过程,其机制可能是按摩能够通过加快清除损伤后自由基的产生,使中性粒细胞浸润减少,降低PLA2活性来保护细胞结构完整性,减轻疼痛。     

不同变性条件下肌酸激酶活力和构象变化的比较

以盐酸胍,十烷基硫酸钠和溴化十烷基三甲铵为变性剂,测定它们对肌酸激酶（ＣＫ）活力和构象之影响。结果发现：ＣＫ活力的丧失明显早于分子整体构象的变化;Ｃ１０Ｓ和Ｃ１０ＮＭ３对ＣＫ的变性有一定的饱和性;而ＧｕＨＣｌ对ＣＫ的变性则没有;溶液ＰＨ增加时,在一定范围内,ＧｕＨＣｌ,Ｃ１０Ｓ和Ｃ１０ＮＭ３对ＣＫ的变性能力都增加,在ＰＨ变化时,利用ＤＴＮＢ测定ＣＫ内埋巯基的暴露来反映构象的变化是成功的。     

五种苏铁属植物的核形态

报道了苏铁属（Cycas L.)5种植物的染色体数目和核型,除多歧苏铁外,其他种均为首次报道。5个种的体细胞中期染色体核型公式分别为：滇南苏铁C.diannanensis K(2n)=2x=22=2m 4sm 4st 12T;潭清苏铁C.tanqingii K(2n)=2x=22=2m 8sm 2st 10T;多歧苏的Cmultipinnata K(2n)=2x=22=4m 8st 2st 8T;巴兰萨苏铁C.balansae K(2n)=2x=xx=2m 4sm 6st 10T。石山苏铁C.miquelii K(2n)=22=2m 6sm(1SAT) 4st 10T;核型均属于3B型。本研究结果支持苏铁属植物的核型从不对称进化的观点;同时,支持将巴兰萨苏铁和石山苏铁归入攀枝花苏铁组的台湾苏铁亚组的观点。     

水分亏缺对番茄叶片气孔特性及叶绿体超微结构的影响

以'辽园多丽'番茄为材料,在塑料大棚内设置每桶2 000(CK)、1 500(Ti),1 000(Tz)、506(T3)和300 mL(T4)5个灌水量处理,利用扫描和透射电镜观察了处理结束时和恢复灌水10 d后番茄叶片的气孔特性和叶绿体的超微结构.结果表明:(1)番茄叶片的总气孔密度、关闭的气孔数及其与总气孔密度的比值随着水分亏缺程度的增加而增大.气孔器的长、宽和开张度则逐渐变小;(2)恢复灌水10 d后,番茄叶片总气孔密度和关闭气孔数均下降,气孔器开张度增大.T,和T2基本恢复至CK水平.但T3和T4与CK差距仍较大.(3)水分亏缺条件下,叶绿体形状细长.被膜完整度下降,T3和T4处理的被膜解体;单位叶绿体基粒数下降,脂滴和淀粉粒数增加;T3处理的叶绿体片层结构松散弯曲,而T4无完整片层结构;恢复灌水后10 d,轻度水分亏缺处理(T1和T2)的叶片气孔和叶绿体超微结构恢复正常,而严霞水分亏缺(T3和T4)却对番茄叶片光合器官造成不可逆伤害,严重影响其光合作用.     

青藏高原毛茛科3种特有植物核型和进化研究

对青藏高原高山冰缘地区毛茛科３种特有植物的核型进行了分析。它们的核型公式（Ｋ）、染色体相对长度组成（Ｃ．Ｒ．Ｌ．）和核型不对称系数（Ａｓ．Ｋ％）分别为：青藏金莲花Ｔｒｏｌｉｕｓｐｕｍｉｌｕｓｖａｒ．ｔａｎｇｕｔｉｃｕｓ：Ｋ（２ｎ）＝６ｍ＋８ｓｍ（２ＳＡＴ）＋２ｓｔ,Ｃ．Ｒ．Ｌ．＝４Ｌ＋４Ｍ２＋４Ｍ１＋４Ｓ,Ａｓ．Ｋ％＝６３．５７,核型属２Ｂ型;甘青乌头Ａｃｏｎｉｔｕｍｔａｎｇｕｔｉｃｕｍ为Ｋ（２ｎ）＝６ｍ＋１０ｓｍ,Ｃ．Ｒ．Ｌ．＝４Ｌ＋８Ｍ１＋４Ｓ,Ａｓ．Ｋ％＝６２．５４,２Ｂ型;单花翠雀花Ｄｅｌｐｈｉｎｉｕｍｃａｎｄｅｌａｂｒｕｍｖａｒ．ｍｏｎａｎｔｈｕｍ为Ｋ（２ｎ）＝６ｍ＋８ｓｍ＋２ｓｔ,Ｃ．Ｒ．Ｌ．＝４Ｌ＋４Ｍ２＋４Ｍ１＋６Ｓ,Ａｓ．Ｋ％＝６４．３４,属３Ｂ型。经同相关近缘种核型资料比较,青藏金莲花核型不对称性和进化程度比金莲花Ｔ．ｃｈｉｎｅｎｓｉｓ低;甘青乌头的核型不对称性和进化程度在其近缘类群（乌头组Ｓｅｃｔ．Ａｃｏｎｉｔｕｍ）已报道的种之内最低;单花翠雀花核型不对称性和进化水平比翠雀组（Ｓｅｃｔ．Ｄｅｌｐｈｉｎａｓｔｒｕｍ）已报道的展毛翠雀花Ｄ．ｋａｍａｏｅｎｓｅｖａｒ．ｇｌａｂｒｅｓｃｅｎｓ、     

Wild-type NM23-H1, but not its S120 mutants, suppresses desensitization of muscarinic potassium current

NM23 (NDP kinase) modulates the gating of muscarinic K+ channels by agonists through a mechanism distinct from GTP regeneration. To better define the function of NM23 in this pathway and to identify sites in NM23 that are important for its role in muscarinic K+ channel function, we utilized MDA-MB-435 human breast carcinoma cells that express low levels of NM23-H1. M2 muscarinic receptors and GIRK1/GIRK4 channel subunits were co-expressed in cells stably transfected with vector only (control), wild-type NM23-H1, or several NM23-H1 mutants. Lysates from all cell lines tested exhibit comparable nucleoside diphosphate (NDP) kinase activity. Whole cell patch clamp recordings revealed a substantial reduction of the acute desensitization of muscarinic K+ currents in cells overexpressing NM23-H1. The mutants NM23-H1P96S and NM23-H1S44A resembled wild-type NM23-H1 in their ability to reduce desensitization. In contrast, mutants NM23-H1S120G and NM23-H1S120A completely abolished the effect of NM23-H1 on desensitization of muscarinic K+ currents. Furthermore, NM23-H1S120G potentiated acute desensitization, indicating that this mutant retains the ability to interact with the muscarinic pathway, but has properties antithetical to those of the wild-type protein. We conclude that NM23 acts as a suppressor of the processes leading to the desensitization of muscarinic K+ currents, and that Ser-120 is essential for its actions.     

Design and Evaluation of Self-Emulsifying Drug Delivery Systems (SEDDS) of Nimodipine

The ability of self-emulsifying drug delivery systems (SEDDS) to improve solubility, dissolution rate and bioavailability of a poorly water-soluble calcium channel blocker, nimodipine (NM) was evaluated in the present investigation. Solubility of NM in various oils, surfactants and cosurfactants was determined. The influence of the ratio of oil to surfactant + cosurfactant, pH of aqueous phase on mean globule size of resulting emulsions was studied by means of photon correlation spectroscopy. The NM loaded SEDDS selected for the in vitro and in vivo studies exhibited globule size less than 180 nm. In vitro dissolution studies indicated that NM loaded SEDDS could release complete amount of NM irrespective of the pH of the dissolution media. Pharmacokinetics of NM suspension, NM oily solution, NM micellar solution and NM SEDDS were evaluated and compared in rabbits. Relative bioavailability of NM in SEDDS was significantly higher than all the other formulations. NM loaded SEDDS were subjected to various conditions of storage as per ICH guidelines for 3 months. NM SEDDS successfully withstood the stability testing.     

Surfactant-induced refolding of lysozyme

The surfactant-lysozyme interaction was investigated by circular dichroism, fluorescence, UV, dynamic light scattering, surface tension, turbidity measurements and lysozyme activity assay. A new way of refolding of lysozyme was found. It was shown that the lysozyme unfolded by anionic surfactants could be renatured by adding cationic surfactants. That is, lysozyme formed precipitate with anionic surfactants, the precipitates could be dissolved by adding a cationic surfactant solution, and then the lysozyme was refolded to its native state spontaneously. Different couples of anionic surfactants and cationic surfactants including C10SO3/C10NE, C12SO3/C10NE, C10SO3/C12NE, C10SO3/C12NB, C10SO4/C10NE and C12SO4/C10NE (C(n)SO3, C(n)SO4, C(n)NE and C(n)NB represent sodium alkyl sulfonate/sulfate, alkyl triethyl/butyl ammonium bromide respectively) were investigated, all of them gave similar results. The results were explained in terms of the differences between the interaction of anionic-cationic surfactants and that of surfactant-lysozyme. It was thought that the formation of mixed micelles of anionic-cationic surfactants is a more favorable process than that of lysozyme-surfactant complexes, which induces the dissociation of lysozyme-surfactant complexes when cationic surfactants were added.     

Interaction of cisplatin with methionine- and histidine-containing peptides: competition between backbone binding,macrochelation and peptide cleavage

The pH- and time-dependent reaction of cis-[PtCl2(NH3)2] with the methionine- and histidine-containing peptides H-Gly-Met-OH, H-Gly-Gly-Met-OH, Ac-His-Gly-Met-OH, and Ac-His-(Ala)3-Met-OH at 313 K has been investigated by ion-pairing reverse phase HPLC and NMR spectroscopy. For equimolar solutions (c=0.8 mM, pH approximately equals 3 or 8.8), initial formation of the kinetically favored S-bound complex is followed by relatively rapid metallation of the neighboring methionine amide nitrogen NM to afford a kappa2NM,S six-membered chelate. The strong trans effect of the methionine S then favors facile NH3 substitution, leading to generation of tridentate complexes such as [Pt(H-Gly-MetH(-1)-OH)-kappa3NG,NM,S)(NH3)]+ or [Pt(H-Ac-His-GlyH(-1)-MetH(-1)-OH-kappa3NG,NM,S)(NH3)]. In the case of H-Gly-Gly-Met-OH, this reaction is accompanied by loss of a second NH3 ligand in alkaline solution to generate the tetradentate kappa4NG1,NG2,NM,S species. In contrast, cleavage of the backbone C(O)-N bond to the second metallated amide nitrogen after t>100 h is common to the tridentate complexes of the tri- and pentapeptides at pH<5. Although an imidazole-coordinated kappa2N3H,S macrochelate is formed throughout the whole range 2.5 < or = pH < or = 10 for Ac-His-Gly-Met-OH, it slowly decays (t=10-1000 h) to the thermodynamically more stable tridentate kappa3NG,NM,S complex. All major final products were separated and fully characterized by NMR and MS.     

硫素营养水平对产油尖状栅藻光合生理及生化组成的影响

以产油尖状栅藻(Scenedesmus acuminatus)为实验材料, 在持续300 μmol photons/(m2·s)光照条件下, 选用3种不同初始Na₂SO₄浓度(2.0S、1.0S对照、0.25S)的改良BG-11培养基, 在Φ3.0 cm×60 cm光生物反应器中进行通气培养, 研究分析硫素营养水平与尖状栅藻产油过程光合生理和生化组成的关系。实验结果表明, 初始硫素浓度对尖状栅藻生长有显著的影响(P<0.05), Na₂SO₄初始浓度为2.0S实验组的生物量最高, 为7.47 g/L, 显著高于1.0S组(6.43 g/L)和0.25S组(4.17 g/L)(P<0.05), 说明加富硫素营养可促进藻细胞的生长。尖状栅藻细胞的叶绿素a、b以及总类胡萝卜素含量变化均与培养基中初始硫素营养水平呈正相关。在培养初期低硫营养有利于藻细胞快速积累碳水化合物, 0.25S实验组碳水化合物含量最高, 占干重的44.37%, 比1.0S和2.0S组分别高出14.43%和13.78%, 培养后期总碳水化合物和蛋白含量均发生不同程度的降低, 转向大量累积油脂, 0.25S实验组的总脂含量最高, 达55.15% DW, 显著高于1.0S和2.0S组(P<0.05)。藻细胞的光合放氧速率、PSⅡ最大光能转化效率(F_v/F_m)、实际光能转换效率(Yield)以及相对电子传递效率(ETR)均与培养液的初始硫素浓度呈正相关, 在整个培养周期中呈先上升后下降的趋势。77 K低温荧光显示, 尖状栅藻在培养初期2个光系统之间存在光能调配现象。上述结果说明, 尖状栅藻细胞的生长、油脂积累和光合生理状况与硫素营养水平直接相关。