A cavity-containing mutant of T4 lysozyme is stabilized by buried benzene.

The hydrophobic cores of proteins are generally well packed, with few cavities. Mutations in which a bulky buried residue such as leucine or phenylalanine is replaced with a small residue such as alanine can create cavities in the core of a protein (our unpublished results). The sizes and shapes of such cavities can vary substantially depending on factors such as local geometry, whether or not a cavity already exists at the site of substitution, and the degree to which the protein structure relaxes to occupy the space vacated by the substituted residue. We show by crystallographic and thermodynamic analysis that the cavity created by the replacement Leu 99----Ala in T4 lysozyme is large enough to bind benzene and that ligand binding increases the melting temperature of the protein by 6.0 degrees C at pH 3.0. Benzene does not, however, bind to the cavity created by the Phe 153----Ala replacement. The results show that cavities can be engineered in proteins and suggest that such cavities might be tailored to bind specific ligands. The binding of benzene at an internal site 7 A from the molecular surface also illustrates the dynamic nature of proteins, even in crystals.     

Solution structure of a minor and transiently formed state of a T4 lysozyme mutant

Proteins are inherently plastic molecules, whose function often critically depends on excursions between different molecular conformations (conformers). However, a rigorous understanding of the relation between a protein's structure, dynamics and function remains elusive. This is because many of the conformers on its energy landscape are only transiently formed and marginally populated (less than a few per cent of the total number of molecules), so that they cannot be individually characterized by most biophysical tools. Here we study a lysozyme mutant from phage T4 that binds hydrophobic molecules and populates an excited state transiently (about 1?ms) to about 3% at 25?°C (ref. 5). We show that such binding occurs only via the ground state, and present the atomic-level model of the 'invisible', excited state obtained using a combined strategy of relaxation-dispersion NMR (ref. 6) and CS-Rosetta model building that rationalizes this observation. The model was tested using structure-based design calculations identifying point mutants predicted to stabilize the excited state relative to the ground state. In this way a pair of mutations were introduced, inverting the relative populations of the ground and excited states and altering function. Our results suggest a mechanism for the evolution of a protein's function by changing the delicate balance between the states on its energy landscape. More generally, they show that our approach can generate and validate models of excited protein states.     

Contributions of hydrogen bonds of Thr 157 to the thermodynamic stability of phage T4 lysozyme

Measurements of changes in structure and stability caused by 13 different substitutions for threonine 157 in phage T4 lysozyme show that the most stable lysozyme variants contain hydrogen bonds analogous to those in the wild-type enzyme and that structural adjustments allow the protein to be surprisingly tolerant of amino-acid substitutions.     

直觉I-fuzzy拓扑空间中,T3,T4分离公理

目的讨论直觉I-fuzzy拓扑空间中的T_3,T_4分离公理以及与T_1,T_2分离公理的关系。方法 L*-格值上Lukasiewicz蕴含算子。结果与结论首先给出直觉I-fuzzy拓扑空间中T_3,T_4分离性的概念,接着得到它们的等价命题,最后讨论了T_3,T_4分离性与T_1,T_2分离性的关系。     

搅拌槽内新型疏水缔合聚合物AP-P4的溶解性

由于疏水缔合聚合物聚丙烯酰胺(AP-P4)的溶解性差,制约了聚合物驱油技术在海上油田的采收率。考察了温度、矿化度、转速、搅拌器型式及搅拌方式对AP-P4溶解时间和聚合物溶液黏度特性的影响,结果表明:温度与转速的提高有利于聚合物的加速溶解,但不利于聚合物溶液的增黏,存在一个适宜聚合物溶解的最佳矿化度范围;剪切力较小、循环量较大的搅拌器更适合AP-P4的溶解;采用翼型搅拌器向上排出流体,先高转速后低转速的搅拌方式能够加快AP-P4的溶解并获得较高的溶液黏度。     

Light-induced hormone conversion of T4 to T3 regulates photoperiodic response of gonads in birds

Reproduction of many temperate zone birds is under photoperiodic control. The Japanese quail is an excellent model for studying the mechanism of photoperiodic time measurement because of its distinct and marked response to changing photoperiods. Studies on this animal have suggested that the mediobasal hypothalamus (MBH) is an important centre controlling photoperiodic time measurement. Here we report that expression in the MBH of the gene encoding type 2 iodothyronine deiodinase (Dio2), which catalyses the intracellular deiodination of thyroxine (T4) prohormone to the active 3,5,3'-triiodothyronine (T3), is induced by light in Japanese quail. Intracerebroventricular administration of T3 mimics the photoperiodic response, whereas the Dio2 inhibitor iopanoic acid prevents gonadal growth. These findings demonstrate that light-induced Dio2 expression in the MBH may be involved in the photoperiodic response of gonads in Japanese quail.     

Acquisition of specific cytotoxic activity by human T4+ T lymphocytes in culture

Mature human T lymphocytes can be separated by monoclonal antibodies OKT4 and OKT8 according to their surface phenotypes into T4+T8- and T4-T8+ subsets. From short-term experiments using bulk cultures, the helper/inducer function has been assigned to the T4+T8- subset and the cytotoxic/suppressor function to the T4-T8+ subset. Thus if T lymphocytes are separated after stimulation in a mixed lymphocyte reaction (MLR), the entire cytotoxic activity is found in the T4-T8+ fraction whereas the T4+T8- fraction shows no detectable cytotoxicity. If, however, T lymphocytes are cloned after MLR and grown in long-term culture, a surprisingly large fraction of T4+ T lymphocyte clones (TLC) shows cytotoxic activity. Here we report that T4+ TLC can acquire specific cytotoxicity during in vitro cultivation.     

Demonstration by NMR of folding domains in lysozyme

Although there has been much speculation on the pathways of protein folding, only recently have experimental data on the topic been available. The study of proteins under conditions where species intermediate between the fully folded and unfolded states are stable has provided important information, for example about the disulphide intermediates in BPTI, cis/trans proline isomers of RNase A3 and the molten globule state of alpha-lactalbumin. An alternative approach to investigating folding pathways has involved detection and characterization of transient conformers in refolding studies using stopped-flow methods coupled with NMR measurements of hydrogen exchange. The formation of intermediate structures has been detected in the early stages of folding of cytochrome c, RNaseA and barnase. For alpha-lactalbumin, hydrogen exchange kinetics monitored by NMR proved to be crucial for identifying native-like structural features in the stable molten globule state. An analogous partially folded protein stable under equilibrium conditions has not been observed for the structurally homologous protein hen egg-white lysozyme, although there is evidence that a similar but transient state is formed during refolding. Here we describe NMR experiments based on competition between hydrogen exchange and the refolding process which not only support the existence of such a transient species for lysozyme, but enable its structural characteristics to be defined. The results indicate that the two structural domains of lysozyme are distinct folding domains, in that they differ significantly in the extent to which compact, probably native-like, structure is present in the early stages of folding.     

含多参数的不确定线性时滞系统的鲁棒镇定

应用李雅普诺夫稳定性理论,给出了一类含多参数的参数不确定系统的鲁棒控制器的一种设计方法．通过构造状态反馈矩阵K,使其满足所给的两个条件,以此得到一族鲁棒稳定控制器．最后以数值实例说明了该设计方法的有效性．文中实例表明,该结论具有更小的保守性．     

Three-dimensional solution structure of w-conotoxin SO3 determined by ~1H NMR

Cone snails (Conus) elaborate a series of conotoxin (CTX) peptides in their venoms to paralyze their prey. Many of these toxins specifically block ion channels in vertebrate and invertebrate neuromuscular systems[1]. The pattern of Cys residues is quite conserved in most CTXs and can be arranged in three major frameworks: CCCC (a-CTX), CCCCCC (m-CTX), CCCCCC (w-CTX). These peptides have been used as important tools in receptor and ion channel studies[2]. Among these toxins, w-CT…     

Massive infection and loss of memory CD4+ T cells in multiple tissues during acute SIV infection

It has recently been established that both acute human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) infections are accompanied by a dramatic and selective loss of memory CD4+ T cells predominantly from the mucosal surfaces. The mechanism underlying this depletion of memory CD4+ T cells (that is, T-helper cells specific to previously encountered pathogens) has not been defined. Using highly sensitive, quantitative polymerase chain reaction together with precise sorting of different subsets of CD4+ T cells in various tissues, we show that this loss is explained by a massive infection of memory CD4+ T cells by the virus. Specifically, 30-60% of CD4+ memory T cells throughout the body are infected by SIV at the peak of infection, and most of these infected cells disappear within four days. Furthermore, our data demonstrate that the depletion of memory CD4+ T cells occurs to a similar extent in all tissues. As a consequence, over one-half of all memory CD4+ T cells in SIV-infected macaques are destroyed directly by viral infection during the acute phase-an insult that certainly heralds subsequent immunodeficiency. Our findings point to the importance of reducing the cell-associated viral load during acute infection through therapeutic or vaccination strategies.     

铬黑T还原法测定铬酸根

本文利用CrO_4~(2-)在0.1～0.2mol/L HCl溶液中氧化铬黑T,用缓冲溶液(NH_3-NH_4Cl)将溶液调到pH≈9.6后,在615nm下测定吸光度的减少,建立了测定Cr(Ⅵ)的新方法.方法的定量检出限为6μg/25ml,CrO_4~(2-)线性范围为0～24和24～48μg/25ml CrO_4~(2-).同时试验了35种共存离子的干扰,对金属离子的干扰消除,可用DDTC CHCl_3萃取分离和加柠檬酸,酒石酸钠掩蔽.此方法用于废水中CrO_4~(2-)的测定,得到满意的结果.     

多时间粒度下时态模式的T3NF分解算法在Delphi中的实现

针对时态数据库中有效地进行数据库设计而提出的通过分析TFD集所具有的良好特性,给出满足时态第三范式（T3NF）的无损分解的多项式时间的算法。本文详细介绍了如何在Delphi开发环境下对此算法进行实现,并给出了与此算法相关的一些具体设计方案。     

木犀草素抑制3T3-L1前脂肪细胞的分化

为研究木犀草素在3T3-L1前脂肪细胞成脂分化过程中的作用,文章探讨木犀草素抑制脂质沉积的作用机制,选取3T3-L1前脂肪细胞作为研究对象,在其分化过程中添加木犀草素,利用噻唑蓝(MTT)实验研究木犀草素对3T3-L1增殖的作用;利用油红O染色确定其对3T3-L1前脂肪细胞脂肪化的影响;利用定量聚合酶链式反应(polymerase chain reaction,PCR)检测木犀草素对3T3-L1脂肪化相关基因的表达影响。结果表明,20μmol/L的木犀草素可以降低3T3-L1细胞的脂肪化,减少脂质聚集,并且降低了3T3-L1细胞中脂肪化相关基因Pparγ、C/EBPα、Ap2和Fas等基因的表达。     

Ce(NO3)3对不同细胞DNA的损伤作用研究

以体外培养３Ｔ３ 细胞和ｌｏｖｏ 细胞为研究对象, 运用单细胞凝胶电泳技术, 探讨了Ｃｅ（ＮＯ３）３ 对这２ 种细胞ＤＮＡ 的损伤作用．结果表明Ｃｅ（ＮＯ３）３ 对２ 种细胞均有损伤作用： 低浓度（１ ｍｍｏｌ／Ｌ） 时,ｌｏｖｏ 细胞的ＤＮＡ 损伤率要明显高于３Ｔ３ 细胞ＤＮＡ 的损伤率, 高浓度（５ｍｍｏｌ／Ｌ） 时, 则２ 种细胞的损伤率几近相同, 提示较低浓度Ｃｅ （ＮＯ３） 对正常细胞和癌细胞存在着不同的作用, 稀土对癌细胞具有选择性杀伤作用; Ｃｅ （ＮＯ３） 具有一定的遗传毒性     

FAAS法直接测定白酒中锰

用 FAAS法直接测定白酒中锰 ,精密度高、准确度好、快速经济、有利于批量样品的测定     

Unusual intron in the immunoglobulin domain of the newly isolated murine CD4 (L3T4) gene

The T-cell surface glycoprotein, CD4, is expressed predominantly on helper T cells and is thought to play a major role in cell-cell interactions. Monoclonal antibodies against CD4 have been shown to block numerous T-cell functions; moreover, recent results suggest that the CD4 molecule may be involved in transmembrane signal transduction. The human CD4 glycoprotein has also been shown to form at least part of the receptor for the AIDS virus, HIV-1. Elucidation of the functions of CD4 will be facilitated by the ability to manipulate the protein by genetic means. Because the mouse system is well suited for a variety of functional studies, we have isolated, sequenced and expressed cDNA clones encoding the murine CD4 (L3T4) glycoprotein. Comparison of the mouse and human CD4 sequences reveals striking evolutionary conservation of the cytoplasmic domain, suggesting that this region is essential for CD4 function. In addition, both the human and mouse CD4 gene contain a large intron in the coding region of the V-like domain. As no other members of the immunoglobulin gene superfamily have been shown to contain similarly placed introns, this finding may have important implications regarding the evolution of this gene family in particular and of introns in general.     

Transformation of NIH 3T3 cells by a human c-sis cDNA clone

The mechanism of leukaemogenic transformation by human T-cell leukaemia/lymphoma virus (HTLV), a retrovirus implicated in the aetiology of certain adult T-cell leukaemias and lymphomas, is unknown but is conceivably associated with the expression of the cellular analogues of retroviral oncogenes. The HUT-102 cell line, derived from a cutaneous T-cell lymphoma and infected with HTLV, expresses several cellular oncogenes. It is unusual among haemopoietic cell lines in that one of these is c-sis, the gene from which the oncogene v-sis of the simian sarcoma virus was derived, and perhaps the gene for platelet-derived growth factor (PDGF). To explore the possible role of c-sis expression in HTLV-induced disease, we have obtained cDNA clones of c-sis from HUT-102 cells. Here we describe two such clones and report that one of them transforms NIH-3T3 cells. This is the first example of transformation of NIH-3T3 cells by a human onc gene other than c-ras or Blym, as well as the first demonstration of transformation by a human cDNA clone.