Proteases from human immunodeficiency virus and avian myeloblastosis virus show distinct specificities in hydrolysis of multidomain protein substrates.

The virally encoded proteases from human immunodeficiency virus (HIV) and avian myeloblastosis virus (AMV) have been compared relative to their ability to hydrolyze a variant of the three-domain Pseudomonas exotoxin, PE66. This exotoxin derivative, missing domain I and referred to as LysPE40, is made up of a 13-kilodalton NH2-terminal translocation domain II connected by a segment of 40 amino acids to enzyme domain III of the toxin, a 23-kilodalton ADP-ribosyltransferase. HIV protease hydrolyzes two peptide bonds in LysPE40, a Leu-Leu bond in the interdomain region and a Leu-Ala bond in a nonstructured region three residues in from the NH2-terminus. Neither of these sites is cleaved by the AMV enzyme; hydrolysis occurs, instead, at an Asp-Val bond in another part of the interdomain segment and at a Leu-Thr bond in the NH2-terminal region of domain II. Synthetic peptides corresponding to these cleavage sites are hydrolyzed by the individual proteases with the same specificity displayed toward the protein substrate. Peptide substrates for one protease are neither substrates nor competitive inhibitors for the other. A potent inhibitor of HIV type 1 protease was more than 3 orders of magnitude less active toward the AMV enzyme. These results suggest that although the crystallographic models of Rous sarcoma virus protease (an enzyme nearly identical to the AMV enzyme) and HIV type 1 protease show a high degree of similarity, there exist structural differences between these retroviral proteases that are clearly reflected by their kinetic properties.     

Concerted generation of Ig isotype diversity in human fetal bone marrow

The human fetal bone marrow B cell compartment of 14- to 21-wk gestational age was examined phenotypically and with respect to Ig H chain commitment and diversity. A dramatic expansion of fetal marrow B cell pools at 16- to 18-wk gestational age characterizes a rapid and concerted chain of differentiation events. Transiently up to 1/4 of nucleated marrow cells are CD20+/CD21+ cells which begin to express surface Ig other than IgM. Limiting dilution analysis of EBV-infected marrow cells delineated a virtually exclusive commitment to IgM production until 15 wk and the absolute and relative number of these cells were small (approximately 5% of comparable adult values). In parallel to the rapid increase in total B cell pools size, cells committed and able to secrete any of the five Ig isotypes are generated by 16-wk gestational age and by 18 wk the frequencies of these cells rapidly reach levels typical for adult peripheral tissue such as blood or lymph node. Fetal L chain diversity always anticipated that observed in adult serum. In addition to rising pool sizes and diverse IgH expression, EBV transformability is a major variable during this period of B cell development with up to 2/3 of B lineage cells transformable, about half of which are pre-B cells. By 21-wk gestational age transformable pre-B cells have disappeared and (as in adult tissue) approximately 10 to 20% of CD20+ cells are transformable. The rapid, concerted expression of full H chain diversity during a narrow period in fetal development is unique to marrow and implies a lymphopoietic process in a privileged site rather than an immunologic differentiation event. During this event, the relative proportions between the different IgH classes expressed, resembled that found in adult tissue, perhaps suggesting that B cell inherent programming rather than only antigenic forces determine heavy chain choice. The staggered expression, early in postnatal life, of IgH regions 3' of the C mu locus may reflect regulatory functions rather than inherent immaturity of the B lineage.     

甘蔗叶不同部位ATP酶活性细胞化学定位

甘蔗叶片,叶鞘和肥厚带韧皮部 ATP 酶活性定位于筛管、伴胞的质膜、内质网和某些伴胞细胞基质、小囊泡和发育成熟的液泡上;叶片韧皮部薄壁细胞、厚壁细胞和厚壁通道细胞质膜及小囊泡中亦显示有 ATP 水解产物;维管束鞘细咆与厚壁细胞或厚壁通道细胞所构成的细胞间隙上也存在有 ATP 酶活性反应产物沉淀。甘蔗叶片大、中、小三种维管束,从小维管束到大维管束,面向细胞间隙的细胞表面上的 ATP 酶活性逐渐增强,而维管束鞘细胞质膜上的 ATP 酶活性则趋于减弱;同一维管束内则以韧皮部细胞的 ATP 酶活性最强。维管束鞘细胞与叶肉细胞之间存在很多的胞间连丝,并表现出高的 ATP 酶活性。讨论了 ATP 酶活性的分布状态与叶肉细胞的光合产物向韧皮部运输的关系。     

聚合酶链式反应检测结核杆菌的研究

以人型结核杆菌基因组DNA为模板,合成二段引物各20个碱基进行聚合酶链式反应(PCR)。经琼脂糖凝胶电泳证实,获得一条245bp扩增带。PCR检测的敏感性染色体基因组DNA为1pg,菌悬液为13个活菌／ml。在特异性试验中,人型结核杆趋,牛型结核杆菌、BCG可见此扩增带。被试的其它14种扰酸菌以及变铅青链霉菌、大肠杆菌质粒Puc19、星状诺卡氏菌、红球菌均未见该扩增带。54例肺结核痰标本3种方法检查的阳性率分别为：萋尼氏抗酸染色16．7％,培养法14．8％,PCR 37．0％。前2种检查方法分别与PCR比较,经统计学处理均有显著性差异(P<0．01)。12例非结核性肺部疾患痰标本抗酸染色和PCR均为阴性。结果表明,PCR技术是快速、敏感、特异诊断结核病的方法。     

Minimal sequence requirement of thrombin receptor agonist peptide.

A site-specific proteolytically generated neoamino terminus of the thrombin receptor having a sequence SFLLRNPNDKYEPF- has been reported to be a functional ligand of the receptor. This discovery raises question on the precise structural requirements of the "tethered ligand" responsible for receptor activation and signal transduction. By examining the agonist activity of a panel of synthetic sequence analogues of thrombin receptor agonist peptides (TRAP) on human platelet aggregation, we determined that the minimal sequence of the human platelet thrombin receptor ligand is SFLL-amide (TRAP1-4, EC50 = 300 uM). An extension of TRAP1-4 by an additional Arg-Asn segment yielded the most potent agonist among the series (TRAP1-6, EC50 = 1.3 microM). Based on the structure-activity relationships, we hypothesized a model of the ligand-binding site of the human platelet thrombin receptor that accommodates a hexapeptide structure. TRAP1-6, when administered intravenously, induced marked intravascular platelet aggregation in the anesthetized guinea pigs.     

Preliminary study of the distribution of the toxic elements As,Cd, and Hg in human hair and tissues by RNAA

In order to study the relationships between trace element concentrations of hair and internal body burdens, a radiochemical NAA technique has been used for determination of the elements As, Cd, and Hg in autopsy samples of liver, kidney-cortex, lung, and hair from 24 male persons who died by accident. High significant positive correlations were observed between the As concentration in hair and in kidney-cortex, and between Cd and Zn concentrations in kidney-cortex. The contents of Cd, both for lung and kidney-cortex, were related to the smoking habits of the subjects.

     

家兔脊髓蛛网膜下腔注射乙酰胆碱对血压和心率的影响

将乙酰胆碱(ACh)注入麻醉家兔脊髓蛛网膜下腔,观察其对心血管活动的影响。结果表明:(1)脊髓蛛网膜下腔注射50～100μg ACh可使血压下降,心率减慢;(2)预先由脊髓蛛网膜下腔注射阿托品,可阻断ACh引起的降压和降心率作用;(3)脊髓蛛网膜下腔注射六甲双铵、酚妥拉明或心得安均不能阻断上述ACh的心血管反应;(4)切断两侧颈部迷走神经,ACh不再使心率减慢,但其降低血压的作用不受到任何影响。 脊髓中ACh水平升高可通过激活胆碱能M-受体引起血压下降和心率减慢。ACh的这种降压作用既没有中枢肾上腺素能受体活动参与,也不是通过迷走神经实现的,可能是由于脊髓交感血管中枢紧张性降低所造成的。     

辽西半干旱地区春小麦农田水分循环特征的研究

一、研究地区的生态条件概述辽西属暖温带半干旱低山丘陵区,自然条件差,干旱少雨,干燥度为1.2左右,年均降水量为480.1mm,可能蒸散量为551.7mm,水分亏缺严重,而且70—75%的雨水降子6—8月份,春季偏少。舂季气温回升快,风速大,土壤水分蒸发损失严重,因此春旱频繁,并经常     

小麦×玉米杂交后代的蛋白质及酯酶同工酶分析

以８ 个普通小麦（Ｔｒｉｔｉｃｕｍ ａｅｓｔｉｖｕｍ Ｌ．）品种为母本,２ 个栽培玉米（Ｚｅａ ｍ ａｙｓ Ｌ．）品种为父本杂交所获得的Ｆ２ 代在形态上出现了明显变异。对其籽粒进行蛋白质电泳分析,得到了如下主要结果：杂交后代的蛋白质谱带较母本有了很大的变异,主要集中在高分子量麦谷蛋白（ＨＭＷ－Ｇｌｕ）区域。杂交后代的蛋白质谱带由５ 种类型构成：１．母本型,占全部测试籽粒的２２．６％ ;２．附加型,占１４．３％ ;３．互补型,占１５．５％ ;４．杂种型,占３０．９％ ;５．缺失型,占１６．７％ 。对“矮杆早”ד紫粘”的Ｆ２ 代籽粒进行酯酶同工酶电泳分析发现,变异主要发生在ＥＳＴ－１ 区。由此看来,小麦×玉米的杂合子中玉米染色体在被排除前后,可以诱发小麦染色体组发生遗传变异     

Human neurotrophin-3: A one-step peptide mapping method and complete disulfide characterization of the recombinant protein

Human neurotrophin-3 (NT-3) is a member of the nerve growth factor (NGF) family of neurotrophic factors, and the recombinant protein is being developed as a therapeutic for neurodegenerative diseases. The final product purity and lot-to-lot variation are monitored routinely by peptide mapping. However, only the N-terminal region of NT-3 was susceptible to proteolysis under native conditions. Complete digestion required that the protein be chemically modified by reduction and S-alkylation prior to proteolysis. Complete proteolytic degradation of the protein was achieved simply by an intial denaturation of NT-3 in 6 M guanidinium chloride (pH 6) for 2 hr at 37°C, followed by a tenfold dilution with the digestion buffer (0.1 M Tris-HCl, 1 mM CaCl₂ at pH 7.0) and immediate addition of chymotrypsin at 1% by weight. Direct comparison of the peptide map with an identical aliquot that had been reduced and alkylated also allowed the establishment of the cystine linkages present in NT-3: Cys¹⁴ to Cys⁷⁹, Cys⁵⁷ to Cys¹⁰⁸, and Cys⁶⁷ to Cys¹¹⁰. This disulfide structure is homologous to the NGF family of neurotrophic factors.