白细胞介素－2的PEG化

用自制的氨基ＰＥＧ化试剂ｒＩＬ－２进行化学修饰,研究了试剂浓度,溶液ｐＨ,反应时间等与ＰＥｃａ－ｒＩＬ－２产率及ＩＬ－２活性保持之间的关系,建立了一套获得稳定修饰度的ＰＥＧ－ｒＩＬ－２的方法。研究发现,反应时间跟修饰度关系不大;溶液ｐＨ对修饰度有一定的影响,中性ｐＨ以上反应都可进行;而试剂浓度直接决定修饰度的高低,过量越多,修饰度越高,而生物活性保留也越低;但低度修饰,对活性几乎没有影响,可保留活性在９５％左右。     

水稻巯基蛋白酶抑制剂的分子修饰与其对稻瘟病菌的抑制作用

水稻巯基蛋白酶抑制剂（ＣＰＩ）经用二硫苏糖醇,对氯汞苯甲酸和碘乙酸修饰后,对木瓜蛋白酶的抑制活性并无改变;用Ｎ－乙基顺丁烯二酰亚胺与ＣＰＩ反应,可以测出ＣＰＩ分子内有１９个巯基被修饰,被修饰后,抑制活性仍无改变,表明水稻ＣＰＩ的抑制活性不需要巯基参与;应用Ｎ－溴代丁二酰亚胺与ＣＰＩ反应,可测出ＣＰＩ分子内有２个Ｔｒｐ被修饰,修饰后,抑制活性全部丧失,表明Ｔｒｐ是保持抑制活性所必需的基团。水稻巯基蛋白酶抑制剂和丝氨酸蛋白酶抑制剂对稻瘟病菌丝体的生长均有抑制作用,但后者的抑制作用比前者更强,若将两种抑制剂混合使用,则对稻瘟病菌丝体的抑制作用非常强烈;当抑制剂加入量达７２μｇ时,即可产生明显的抑制作用。     

Effects of nucleotide analogs on ATP hydrolysis by P-type ATPases. Comparison between the canine kidney (Na++ K+) ATPase and maize root H+-ATPase

The mechanism by which chemical energy is converted into an electrochemical gradient by P-type ATPase is not completely understood. The effects of ATP analogs on the canine kidney (Na⁺+ K⁺) ATPase were compared to effects of the same analogs on the maize (Zea mays L. cv. W7551) root H⁺-ATPase in order to identify probes for the ATP binding site of the maize root enzyme and to determine potential similarities of ATP hydrolysis mechanisms in these two enzymes. Six compounds able to modify the ATP binding site covalently were compared. These compounds could be classed into three distinct groups based on activity. The first group had little or no effect on catalytic activity of either enzyme and included 7-chloro-4-nitrobenz-2-oxa-1.3-diazole. The second group, which included azido adenine analogs. fluorescein isothiocyanate and 5′-p-fluorosulfonylbenzoyladenine, were inhibitors of ATP hydrolysis by both enzymes. However, the sensitivity of the (Na⁺+ K⁺) ATPase to inhibition was much greater than that exhibited by the maize root enzyme. The third group, which included periodate treated nucleotide derivatives and 2′,3′-o-(4-benzoylbenzoyl)adenosine triphosphate. inhibited both enzymes similarly. This initial screening of these covalent modifiers indicated that 2′,3′-o-(4-benzoylbenzoyl)adenosine triphosphate was the optimal covalent modifier of the ATP binding site of the maize root enzyme. Certain reagents were much more effective against the (Na⁺+ K⁺) ATPase than the maize root enzyme, possibly indicating differences in the ATP binding and hydrolysis pathway for these two enzymes. Two ATP analogs that are not covalent modifiers were also tested: the trinitrophenyl derivatives of adenine nucleotides were better than 5′-adenylylimidodiphosphate for use as an ATP binding probe.     

Amino acid sequence of the minor isomorph of the crustacean hyperglycemic hormone (CHH-II) of the Mexican crayfish Procambarus bouvieri (Ortmann): Presence of a d-amino acid

The primary structure of the neurohormone crustacean hyperglycemic hormone (CHH-II) was determined by means of enzymatic digestions, manual Edman degradation, and mass spectrometry. CHH-II is a 72 residue peptide (molecular mass 8388 Da), with six cysteines forming three disulfide bridges that connect residues 7–43, 23–39, and 26–52. The peptide has blocked N- and C-termini, and lacks tryptophan, histidine, and methionine. The CHH-I and CHH-II of Procambarus bouvieri have identical sequences and elicit levels of hyperglycemia that are not distinguishable. The difference between the two isomorphs consists in a posttranslational modification of a l-Phe in CHH-I to a d-Phe in CHH-II at the third position from the N-terminus.     

Mammalian myristoyl CoA: protein N-myristoyltransferase

Myristoyl CoA:Protein N-myristoyltransferase (NMT) is the enzyme which catalyses the covalent transfer of myristate from myristoyl CoA to the amino-terminal glycine residue of protein substrates. Although NMT is ubiquitous in eukaryotic cells, the enzyme levels and cellular distribution vary among tissues. In this article, we describe the properties of mammalian NMT(s) with reference to subcellular distribution, molecular weights, substrate specificity and the possible involvement of NMT in pathological processes. The cytosolic fraction of bovine brain contains multiple forms of NMT activity whereas bovine spleen contains only a single form. In bovine brain and spleen, the cytosol contained majority of NMT activity. In contrast, rabbit colon and rat liver NMT activity was predominantly particulate. Regional differences in NMT activity have been observed in both rabbit intestine and bovine brain. Results from our laboratory along with the existing knowledge, provide evidence for the existence of tissue specific isozymes of NMT.     

Post-translational modification of proteins and the discovery of new medicine

Post-translational modifications are fundamental to processes controlling behaviour, including cellular signaling, growth and transformation. As the molecular basis of protein modifications in normal and disease processes are becoming better defined, so new strategies for designing therapeutic entities to control complex disease processes are emerging.     

神经生长因子结构与功能研究进展

神经生长因子(NGF)是神经营养因子家族的典型代表, 它控制着脊椎动物周围和中枢神经系统中部分神经元的发育和存活.NGF的三维结构是以“胱氨酸结”和β折叠为基础,它以二聚体的形式结合细胞表面的受体从而发生生物学效应.参与这些反应的氨基酸残基已通过化学修饰和定点突变法加以确定,这有助于更进一步理解其结构与功能的关系.     

Posttranslational Maturation of the Prohormone Convertase SPC3 In Vitro

Abstract: The subtilisin-like prohormone convertase SPC3 is likely to play a role in the biosynthesis of a variety of biologically active peptides. SPC3 undergoes a series of posttranslational processing events during its biosynthesis. Multiple forms have been identified that show varying degrees of truncation at the carboxyl terminus. In this study we show that the 86-kDa form of recombinant SPC3 with an intact carboxyl terminus can undergo rapid carboxyl-terminus truncation to produce a 64-kDa form. We have defined the optimal conditions for carboxyl-terminus truncation in vitro. The carboxyl-terminus truncation reaction was less calcium sensitive, active over a broader pH range, and showed differences in inhibitor sensitivity compared with the enzymatic activities of full-length and truncated forms of SPC3 toward a fluorescent peptide substrate. Increases in enzymatic activity of 86-kDa SPC3 were also measured over a time frame consistent with conversion to the 64-kDa form. However, similar specific activities for both forms of the enzyme suggest such activity increases may not be due to carboxyl-terminus truncation. The different enzymatic properties of the major molecular forms of SPC3 highlight the importance of understanding the molecular events regulating carboxyl-terminal processing of this endoprotease.     

Recent approaches to probe functional groups in ribonuclease P RNA by modification interference

Modification interference is a powerful method to identify important functional groups in RNA molecules. We review here recent developments of techniques to screen for chemical modifications that interfere with (i) binding of(pre-)tRNA to bacterial RNase P RNA or (ii) pre-tRNA cleavage by this ribozyme. For example, two studies have analyzed positions at which a substitution of sulfur for thepro-Rp oxygen affects tRNA binding [1] or catalysis [2]. The results emphasize the functional key role of a central core element present in all known RNase P RNA subunits. The four sulfur substitutions identified in one study [2] to inhibit the catalytic step also interfered with binding of tRNA toE. coli RNase P RNA [1]. This suggests that losses in binding energy due to the modification at these positions affect the enzyme-substrate and the enzyme-transition state complex. In addition, the two studies have revealed, for the first time, sites of direct metal ion coordination in RNase P RNA. The potentials, limitations and interpretational ambiguities of modification interference experiments as well as factors influencing their outcome are discussed.Abbreviations nt nucleotide(s) - PAGE polyacrylamide gel electrophoresis     

Anatomical studies of branchlet abscission related to crown modification in Quercus cerris L.

The basic anatomy of lateral twig insertion onto the main branch in both healthy and damaged Quercus cerris L. trees was studied. An abscission zone is always present: in healthy trees it is formed by a smaller number of cell layers than in damaged ones, where it is more evident with many layers of cells. Cells of the abscission zone are roundish, with many intercellular spaces between them; cell walls are thin, non-lignified and without secondary walls. No starch was found in cells of the abscission zone, where, instead, a few scattered calcium oxalate druses are seen.