tRNase Z: the end is not in sight

Although the enzyme tRNase Z has only recently been isolated, a plethora of data has already been acquired concerning the enzyme. tRNase Z is the endonuclease that catalyzes the removal of the tRNA 3′ trailer, yielding the mature tRNA 3′ end ready for CCA addition and aminoacylation. Another substrate cleaved by tRNase Z is the small chromogenic phosphodiester bis(p-nitrophenyl)phosphate (bpNPP), which is the smallest tRNase Z substrate known so far. Hitherto the biological function as tRNA 3′-end processing enzyme has been shown only in one prokaryotic and one eukaryotic organism, respectively. This review summarizes the present information concerning the two tRNase Z substrates pre-tRNA and bpNPP, as well as the metal requirements of tRNase Z enzymes. Received 29 March 2007; received after revision 15 May 2007; accepted 21 May 2007     

The cytotoxicity of eosinophil cationic protein/ribonuclease 3 on eukaryotic cell lines takes place through its aggregation on the cell membrane

Human eosinophil cationic protein (ECP)/ ribonuclease 3 (RNase 3) is a protein secreted from the secondary granules of activated eosinophils. Specific properties of ECP contribute to its cytotoxic activities associated with defense mechanisms. In this work the ECP cytotoxic activity on eukaryotic cell lines is analyzed. The ECP effects begin with its binding and aggregation to the cell surface, altering the cell membrane permeability and modifying the cell ionic equilibrium. No internalization of the protein is observed. These signals induce cell-specific morphological and biochemical changes such as chromatin condensation, reversion of membrane asymmetry, reactive oxygen species production and activation of caspase-3-like activity and, eventually, cell death. However, the ribonuclease activity component of ECP is not involved in this process as no RNA degradation is observed. In summary, the cytotoxic effect of ECP is attained through a mechanism different from that of other cytotoxic RNases and may be related with the ECP accumulation associated with the inflammatory processes, in which eosinophils are present. Received 26 October 2007; accepted 23 November 2007     

离子交换色谱流动相组成对核糖核酸酶的折叠影响

用离子交换色谱法（IEC）研究了还原变性核糖核酸酶（RNase A）折叠过程中，复性缓冲液种类及pH值、流动相中脲浓度及盐种类对还原变性核糖核酸酶复性的影响。发现pH为弱碱性，用PBS作为缓冲液能提高复性效率；当流动相中含有2．0mol／L脲时复性产率较高；洗脱剂用NaCl时活性回收率较高。     

绿色荧光蛋白融合人核糖核酸酶抑制因子逆转录病毒载体pLNCX-EGFP-C1-hri的构建及鉴定

构建表达重组绿色荧光蛋白融合人核糖核酸酶抑制因子的逆转录病毒载体pLNCX-EGFP-c1-hri,为探讨人核糖核酸酶抑制因子抗肿瘤作用机制打下基础.用亚克隆法,将目的片段egfp-c1-hri从表达载体pEGFP-C1-hri克隆到pLNCX上,用酶切筛选得到阳性克隆后,用脂质体法将其转染到小鼠黑色素瘤细胞B16中,用800 mg/L G418筛选2周后,在荧光显微镜下检测其表达.双酶切鉴定得到pLNCX-EGFP-C1-hri阳性克隆,荧光显微镜显示绿色荧光在B16细胞质中高效表达.成功构建逆转录病毒表达载体pLNCX-EGFP-C1-hri.     

B12基因在小鼠部分肝切除后的表达差异分析

B12是一种受TNF-α诱导的蛋白．利用肝脏部分切除小鼠模型以及核酸酶保护反应研究了B12基因在肝脏部分切除后不同时间的表达差异，发现在受损后28h，B12的表达开始增强，到36h表达一直呈上升趋势，推测该基因可能与肝脏修复过程中DNA的复制有关，为B12基因的进一步的功能研究提供了理论基础．     

Calf thymus ribonuclease H IIa activity lacks ribonuclease H specificity

Summary Less purified fractions of ribonuclease H IIa activity of calf thymus display divalent cation-dependent ribonuclease H activity and divalent cation-independent ribonuclease activity. Because the ratio of the two enzyme activities does not change during successive chromatographic procedures, we suggest that ribonuclease H IIa activity is indeed able to degrade both ssRNA and the RNA moiety of RNA·DNA-hybrids. Ribonuclease H IIa activity can therefore be differentiated from calf thymus ribonuclease H I and H IIb by its lack of ribonuclease H specificity. The native molecular mass of ribonuclease H IIa activity is between 23 and 28 kDa. Under denaturing conditions a 23 kDa-protein band copurifies with the enzyme activity suggesting that this enzyme is monomeric.     

Thermal Transition of Ribonuclease A Observed Using Proton Nuclear Magnetic Resonance

IntroductionBovine pancreatic ribonuclease A ( RNase A) ,asingle- domain protein that has four disulfidebonds,has played a crucial role in studies ofprotein structure,folding and enzyme catalysis.We propose extending these studies by imageanalysis through the use of nuclear magneticresonance ( NMR) spectroscopy,which has theadvantage of direct,continuous,dynamic,nondestructive,and quantitative monitoring.Absorbance measurements at 2 86nm/2 87nm( UV difference spectroscopy) have beentraditi…     

钙延缓黄瓜子叶衰老及其对RNase与活性氧清除有关酶活性的反应

在黄瓜子叶衰老期间外源 Ca~(2+)延缓内源结合 Ca~(2+)和总 Ca~(2+)含量的降低.Ca~(2+)处理5d的子叶,各亚细胞结构部分 RNase 活力均有明显的抑制效应,但不同亚细胞组分的反应程度不尽相同,线粒体对外源 Ca~(2+)处理最敏感,叶绿体则较迟钝,1mMCa~(2+)处理的子叶,SOD 活力高于对照47%.10mMCa~(2+)处理的子叶仅是对照的48%,Ca~(2+)对衰老过程中 SOD 活力作用可能与体内 Ca·CaM 系统有关.子叶衰老过程中过氧化物酶和过氧化氯酶活力均受到 Ca~(2+)的刺激.但 Ca~(2+)对过氧化氢酶活力刺激效应比过氧物酶小得多.上述结果表明,Ca~(2+)的作用可能与体内自由基清除有关的酶有关;外源 Ca~(2+)对暗诱导黄瓜子叶过氧化物酶作用通过释放胞壁以离子键结合的酶来完成.     

核糖核酸酶抑制因子的制备及急性毒性实验

对新鲜人胎盘进行硫酸铁分级沉淀，透析，DEAE-纤维素-23离子交换层析，制取初步纯化的核糖核酸酶抑制因子（RI）．小鼠急性毒性实验结果表明，RI对小鼠没有急性毒性．     

Introduction: the ribonuclease A superfamily

In this multi-author issue several aspects of the ribonuclease A superfamily are reviewed. This superfamily can be subdivided in a number of mammalian and other vertebrate ribonuclease families. In the introduction chapter the titles of the other contributions are presented. There is little uniformity in the nomenclature of ribonucleases, caused in part by gene duplications, which have occurred independently in several mammalian lineages, and which are nice examples for explaining orthology and paralogy in molecular evolution.