Total chemical synthesis and NMR characterization of the glycopeptide tx5a, a heavily post-translationally modified conotoxin, reveals that the glycan structure is alpha-D-Gal-(1-->3)-alpha-D-GalNAc.

The 13-amino acid glycopeptide tx5a (Gla-Cys-Cys-Gla-Asp-Gly-Trp*-Cys-Cys-Thr*-Ala-Ala-Hyp-OH, where Trp* = 6-bromotryptophan and Thr* = Gal-GalNAc-threonine), isolated from Conus textile, causes hyperactivity and spasticity when injected intracerebral ventricularly into mice. It contains nine post-translationally modified residues: four cysteine residues, two gamma-carboxyglutamic acid residues, and one residue each of 6-bromotryptophan, 4-trans-hydroxyproline and glycosylated threonine. The chemical nature of each of these has been determined with the exception of the glycan linkage pattern on threonine and the stereochemistry of the 6-bromotryptophan residue. Previous investigations have demonstrated that tx5a contains a disaccharide composed of N-acetylgalactosamine (GalNAc) and galactose (Gal), but the interresidue linkage was not characterized. We hypothesized that tx5a contained the T-antigen, beta-D-Gal-(1-->3)-alpha-D-GalNAc, one of the most common O-linked glycan structures, identified previously in another Conus glycopeptide, contalukin-G. We therefore utilized the peracetylated form of this glycan attached to Fmoc-threonine in an attempted synthesis. While the result-ing synthetic peptide (Gla-Cys-Cys-Gla-Asp-Gly-Trp*-Cys-Cys-Thr*-Ala-Ala-Hyp-OH, where Trp* =6-bromotryptophan and Thr* = beta-D-Gal-(1-->3)-alpha-D-GalNAc-threonine) and the native peptide had almost identical mass spectra, a comparison of their RP-HPLC chromatograms suggested that the two forms were not identical. Two-dimensional 1H homonuclear and 13C-1H heteronuclear NMR spectroscopy of native tx5a isolated from Conus textile was then used to determine that the glycan present on tx5a indeed is not the aforementioned T-antigen, but rather alpha-D-Gal-(1-->3)-alpha-D-GalNAc.     

Preparation and properties of pure, full-length IclR protein of Escherichia coli. Use of time-of-flight mass spectrometry to investigate the problems encountered.

IclR protein, the repressor of the aceBAK operon of Escherichia coli, has been examined by time-of-flight mass spectrometry, with ionization by matrix assisted laser desorption or by electrospray. The purified protein was found to have a smaller mass than that predicted from the base sequence of the cloned iclR gene. Additional measurements were made on mixtures of peptides derived from IclR by treatment with trypsin and cyanogen bromide. They showed that the amino acid sequence is that predicted from the gene sequence, except that the protein has suffered truncation by removal of the N-terminal eight or, in some cases, nine amino acid residues. The peptide bond whose hydrolysis would remove eight residues is a typical target for the E. coli protease OmpT. We find that, by taking precautions to minimize Omp T proteolysis, or by eliminating it through mutation of the host strain, we can isolate full-length IclR protein (lacking only the N-terminal methionine residue). Full-length IclR is a much better DNA-binding protein than the truncated versions: it binds the aceBAK operator sequence 44-fold more tightly, presumably because of additional contacts that the N-terminal residues make with the DNA. Our experience thus demonstrates the advantages of using mass spectrometry to characterize newly purified proteins produced from cloned genes, especially where proteolysis or other covalent modification is a concern. This technique gives mass spectra from complex peptide mixtures that can be analyzed completely, without any fractionation of the mixtures, by reference to the amino acid sequence inferred from the base sequence of the cloned gene.     

尿素氮-葡萄糖双功能分析仪的研究

由固定化脲酶、谷氨酸脱氢酶、谷氨酸氧化酶、葡萄糖氧化酶的复合酶膜组成的双电极系统,可以同时测定尿素氮和葡萄糖的含量,每次进样量为25μl,20s即可测定出尿素氮和葡萄糖的含量。在0～60mg/dl尿素氮、0～500mg/dl葡萄糖范围内具有良好的线性关系。连续测定20次的变异系数分别为1.02％和1.05％。酶膜使用寿命为两星期以上。此仪器可广泛应用于临床检验和体育训练中。     

二氢叶酸还原酶结合底物的去除

分析了应用氨甲蝶呤（MTX-Agarose）亲和层析法提纯的鸡肝二氢叶酸还原酶的组成和性质.建立了用平面粒度胶等电聚焦法去除与酶紧密结合底物的方法．讨论了结合底物对酶构象研究的影响,并指出,用未完全去除结合底物的酶研究酶在变性过程构象变化会得到错误的结论．     

哺乳动物透明带糖蛋白及其生物活性

透明带在哺乳动物的受精过程中发挥着重要作用,透明带化学成分及其生物活性的研究对于了解精卵识别、顶体反应的诱导等过程的分子机理具有重要意义．近10年来的研究表明透明带主要由糖蛋白（ZPGPs）组成,是在细胞质内合成后转移至透明带的.大多数动物的ZPGPs为4～5种,不同种类动物的ZPGPs的氨基酸序列存在高度同源性.在受精过程中,ZPGPs具有精子受体和诱导顶体反应的双重作用,ZPGPs含有N－连接和O－连接寡聚糖,这些寡糖链是ZPGPs行使生物活性不可缺少的部分．     

具收获与投放的三种群Volterra模型的周期轨道

本文在文〔2〕的基础上,研究三维动力系统的Lyapunov分枝问题．放宽了文〔2〕中关于向量场函数的限制,相应地给出从空间闭轨族扰动产生孤立周期解的判定方法．应用判定定理及椭圆函数的积分技巧,研究了具有收获与投放的三种群Volterra模型,得到了存在周期轨道的充分条件．     

Morphometric Analysis of Hepatocellular carcinoma

Ｍｏｒｐｈｏｍｅｔｒｉｃ　Ａｎａｌｙｓｉｓ　ｏｆ　Ｈｅｐａｔｏｃｅｌｌｕｌａｒ　ｃａｒｃｉｎｏｍａＬａｉＭａｏ－ｄｅ（来茂德）;ＣｈｅｎＰｅｉ－ｈｕｉ（陈培辉）ａｎｄＺｈｏｕＳｈｕｉ－ｙｕｎ（周水云）（ＤｅｐａｒｔｍｅｎｔｏｆＰａｔｈｏｌｏｇｙ,Ｚｈ...     

欧洲伪叶甲Lagria hirta L.种群季节变化和幼虫生长发育(鞘翅目:伪叶甲科)(英)

欧洲伪叶甲Lagria hirta L.野外种群季节变化特点鲜明:成虫仅发生于6月底至8月初,幼虫除6月份外全年可见。基于野外观察和测量幼虫头宽,发现这种甲虫的生殖季节很短,在6至8月份;卵孵化后幼虫生长发育,在越冬前可达第3、4或5龄(个别达6龄);幼虫越冬后,经过总共8—10龄,于6月份化蛹。冬季幼虫发育减慢但不完全停滞(非参数型中数检验,P<0.01)。这项研究改变了以前的观点,即认为该甲虫的幼虫发育只有4或5龄,同时也对这种甲虫的生命周期研究有促进作用。     

在吲哚乙酸不同位点偶联载体蛋白对其抗体特异性的影响

分别选择吲哚乙醇分子上的Ｃ１位羧基和吲哚环上的Ｎ位作为偶联载体蛋白的位点,用混合酸酐法和甲醛搭桥法分别合成了两种免疫原ＩＡＡ—ＣＯ—ＮＨ一ＨＳＡ和ＩＡＡ－Ｎ－ＢＳＡ,并进而制得了对吲哚乙酸侧链识别能力不同的两种多克隆抗体,分别可特异识别甲酯化ＩＡＡ和游离态ＩＡＡ;用碳化二亚胺法和甲醛搭桥法分别合成ＩＡＡ—ＣＯ—ＮＨ－ＢＳＡｂＩＡＡ—Ｎ—ＯＶＡ两种复合物,以之为包被物,建立了两种ＩＡＡＥＬＩＳＡ。其灵敏度分别为０．３５ｐｍｏｌ和１．８０ｐｐｍｏｌ;检测范围分别为０．７８～８００ｐｍｏｌ和１．９５～２０００ｐｍｏｌ;批内变异系数分别为４．４５％和４．７９％;批间变异系数分别为１．１５％和１．５０％。笔者用这两种ＥＬＩＳＡ检测了兰花气生根和桑树苗样品中ＩＡＡ的含量,发现两种检测结果相当一致。