Rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. Identification of essential sulfhydryl residues in the primary sequence of the enzyme

The kinase and sugar phosphate exchange reactions of rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase were inactivated by treatment with 5'-p-fluorosulfonylbenzoyladenosine or 8-azido-ATP, but activity could be restored by the addition of dithiothreitol. This inactivation was accompanied by incorporation of 5'-p-sulfonylbenzoyl[8-14C]adenosine into the enzyme that was not released by the addition of dithiothreitol. The lack of effect of ATP analogs on the ATP/ADP exchange or on bisphosphatase activity and reversal of their effects on the kinase and sugar phosphate reactions by dithiothreitol suggest that 1) they reacted with sulfhydryl groups important for sugar phosphate binding in the kinase reaction, and 2) the inactivation of the kinase by these analogs involves a specific reaction that is not related to their general mechanism of attacking nucleotide-binding sites. In addition, alkylation of the enzymes' sulfhydryls with iodoacetamide prevented inactivation by 5'-p-fluorosulfonylbenzoyladenosine, suggesting that the same thiols were involved. o-Iodosobenzoate inactivated the kinase and sugar phosphate exchange; the inactivation was reversed by dithiothreitol; but there was no effect on the bisphosphatase or nucleotide exchange, indicating that oxidation occurred at the same sulfhydryl that are associated with sugar phosphate binding. ATP or ADP, but not fructose 6-phosphate, protected these groups from modification by 5'-p-fluorosulfonylbenzoyladenosine, 8-azido-ATP, and o-iodosobenzoate. ATP also induced dramatic changes in the circular dichroism spectrum of the enzyme, suggesting that adenine nucleotide protection of thiol groups resulted from changes in enzyme secondary structure. Analysis of cyanogen bromide fragments of 14C-carboxamidomethylated enzyme showed that all radioactivity was associated with cysteinyl residues in a single cyanogen bromide fragment. Three of these cysteinyl residues are clustered in a 38-residue region, which probably plays a role in maintaining the conformation of the kinase sugar phosphate-binding site.     

Purification and characterization of hen oviduct microsomal signal peptidase

Hen oviduct signal peptidase requires only two proteins for proteolysis of fully synthesized secretory precursor proteins in vitro: one with a molecular mass of 19 kilodaltons (kDa) and one which is a glycoprotein whose mass varies from 22 to 24 kDa depending on the extent of glycosylation. Purified signal peptidase has been analyzed both as part of an active catalytic unit and after electroelution of the individual proteins out of a preparative polyacrylamide gel. The multiple forms of the glycoprotein component of signal peptidase bind to concanavalin A and are shown to be derived from the same polypeptide backbone. Removal of their oligosaccharides by digestion with N-glycanase converts these proteins to a single 19.5-kDa polypeptide. The glycoproteins all exhibit very similar profiles following individual digestion with trypsin and separation of the resulting peptides by reverse-phase high-performance liquid chromatography. In addition, sequence analysis of selected peptides from corresponding regions in chromatograms representing each form of the glycoprotein reveals the same amino acid sequences. The 19-kDa signal peptidase protein does not bind concanavalin A, has a distinct tryptic peptide map from that of the glycoprotein, and appears to share no amino acid sequences in common with the glycoprotein. Its copurification on a concanavalin A-Sepharose column indicates that it must interact directly with the glycoprotein subunit.     

Facultative parthenogenesis and sex-ratio evolution

Summary Phenotypic models of selection are used to determine the effect of facultative parthenogenesis on the production of males in a spatially variable environment when (i) sex determination is under strict genetic control, and (ii) when sex may be environmentally determined. The results show that when sex is under strict genetic control and there is some chance of maturing in isolation, selection favors a female-biased sex ratio. When sex can be environmentally induced by cues which indicate high density, selection favors a mixture of genetic and environmental control, such that half the individuals always become female and the other half become females when isolated and become males when not isolated.     

人体单臂间歇运动对发汗调定点的影响

本工作系在微小气候相对恒定条件下,对10名健康男青年每人进行四项实验。实验 Ⅰ 为测定双侧腿足浸入43℃水中,诱发左前臂屈侧显现定量汗点时的口腔温度(舌下)阈值,作为发汗调定点参考值(ToSSP);实验 Ⅱ 为 Ⅰ 附加右臂间歇轻负荷运动(77W)时测定 ToSSP,部分对象还记录了皮肤电反应;实验 Ⅲ、Ⅳ 为 Ⅰ、Ⅱ 均附加4.5m/s 气流(22—25℃)直吹头面部,再分别测定 ToSSP。实验 Ⅰ 与 Ⅱ 同体对照22人次,Ⅲ 与 Ⅳ 同体对照24人次。结果表明,实验 Ⅱ、Ⅳ 的 ToSSP 均值及其潜伏期均值分别较 Ⅰ、Ⅲ 者降低(P<0.01)或缩短(P<0.001);Ⅰ、Ⅱ间的 ToSSP 均值差、潜伏期均值差,分别与 Ⅲ、Ⅳ 之间者无显著差异(P>0.2);Ⅱ、Ⅳ 的ToSSP 均值各与其实验开始前的口温均值亦无明显差异(P>0.5)。此结果支持运动时体温调定点下降的论点,并提示在研究体温调定点活动时,以 ToSSP 为指标较用发汗速率为优越,因 ToSSP 不为许多干扰因素所影响。     

The EH2 reduced intermediate of glutathione reductase contains oxidised flavin-while EH4 does not

Glutathione reductase is a flavoprotein whose x-ray structure has been established. Functional data and the x-ray structure are consistent with a mechanism of reaction in which NADPH reacts with the enzyme to produce a two electron, EH2, and four electron, EH4, intermediate. The former is competent for the transfer of electrons to the substrate glutathione. Several structures are possible for the two NADPH intermediates; in order to aid in the determination of the structure of these intermediates, we have determined their resonance Raman spectra at two excitation frequencies. These studies establish that the EH2 intermediate is an oxidized flavin species while the EH4 species is not. Furthermore, the most likely structure for EH2 involves a charge transfer donation of electrons from the anion of cys-63 to the N5 position of flavin.     

我国恙虫蚴的三个新种(蜱螨目, 恙虫科)

近几年来,我国科学家们在恙虫方面的研究工作巳经取得了显著的成就。然而,这对于拥有辽阔疆土的我国来说,实在只不过是一个开始。我们在这方面的工作也做得很少,虽然从1953年开始收集标本,由于各种条件所限制,未能及时整理,以致大量标本散失或霉烂,十分可惜。本文内容,仅仅是将我站恙虫病调研工作组1957年在本省各地所采集的材料中的三种恙虫蚴加以描述,该三种恙虫蚴均系文献上未曾记载过的种类。     

Synthetic substrate for eukaryotic signal peptidase. Cleavage of a synthetic peptide analog of the precursor region of preproparathyroid hormone

A synthetic peptide analog of the precursor region of preproparathyroid hormone has been shown to be a specific substrate for hen oviduct signal peptidase. The sequence of the 31-residue peptide is Ser-Ala-Lys-Asp-norleucine (Nle)-Val-Lys-Val-Nle-Ile-Val-Nle-Leu-Ala-Ile-Ala-Phe-Leu-Ala-Arg-Ser-As p-Gly-Lys-Ser-Val-Lys-Lys-Arg-D-Tyr-amide (Caulfield, M. P., Duong, L. T., O'Brien, R., Majzoub, J. A., and Rosenblatt, M. (1988) Mol. Endocrinol. 2, 452-458). This sulfur-free signal peptide analog can be labeled with 125I on the C-terminal D-tyrosine and is cleaved by purified hen oviduct signal peptidase between Gly and Lys, the correct site of cleavage of preproparathyroid hormone in vivo. Amino acid sequence analysis of the cleavage product released 125I at the seventh cycle of Edman degradation, confirming that enzymatic cleavage occurs at the physiological site. Synthetic peptide analogs of the substrate with Lys, Pro, or Asp substituted for Nle-18 were poor substrates for the enzyme and were also poor competitive inhibitors of catalysis, suggesting that modifications at position -18, 12 amino acids from the site of cleavage, directly influence binding by the enzyme. Analysis of the reactivity of signal peptidase with these synthetic peptides provides insight into the cleavage specificity requirements of this eukaryotic signal peptidase.     

假菠萝麻蛋白酶分离纯化及特性研究

从假菠萝麻叶汁中用乙醇分部沉淀法得到一种蛋白酶,对酪蛋白有强烈的水解活性,经Ｓｅｐｈａｄｅｘ Ｇ－１００柱层析和ＳＰ－Ｓｅｐｈａｄｅｘ Ｃ－５０离子交换柱层析等步骤纯化,可得到六角形结晶。结晶酶液经ＰＡＧＥ圆盘电泳,每条胶柱上只显示一条蛋白带,其活性在ｐＨ４。５－１０、５５℃范围内较稳定,最适ｐＨ８．５最适温度５０℃,Ｋｍ（酪蛋白）值为０．１８５％（Ｗ／Ｖ）。用ＳＤＳ－ＰＡＧＥ和Ｓｅｐｈａｄｅｘ     

一种新发现的镉超积累植物--钻叶紫菀(Aster subulatus Michx.)

通过野外调查和温室营养液砂培试验,发现并鉴定出钻叶紫菀（Aster subulatus Michx.）是一种新的镉（Cd）超积累植物。调查结果发现,钻叶紫菀对土壤中高含量的Cd有很强的忍耐、吸收和积累能力,其地上部茎、叶Cd含量分别为90.0-150.7mg/kg和119.8-172.6mg/kg,平均值分别为132.8mg/kg和139.2mg/kg。砂基营养液培养试验证明,钻叶紫菀对生长介质中的Cd有很强的忍耐能力,当生长介质中Cd浓度高达150mg/L时,植株仍生长正常,其株高与对照相比无显著差异;地上部Cd含量及其积累量均随生长介质中Cd浓度的增加而增加,当生长介质中Cd浓度为120mg/L时,地上部茎Cd含量和积累量达到最高值,分别为5672.50mg/kg、4.93mg/株。结果表明,钻叶紫菀是一种新的Cd超积累植物,为今后探明植物超积累Cd的机理和Cd污染土壤的植物修复提供一种新的种质资源。