Purification and Characterization of a Substrate-Size-Recognizing Metalloendopeptidase from Streptococcus cremoris H61

During the ripening of Gouda-type cheese, two kinds of endopeptidases were found to participate in the degradation of αs1-CN(f1-23), a specific product from αs1-casein hydrolyzed by chymosin. One of the endopeptidases, lactic acid bacteria endopeptidase (LEP-II), which can recognize the size of its substrates, has already been purified and characterized (T. R. Yan, N. Azuma, S. Kaminogawa, and K. Yamauchi, Eur. J. Biochem. 163:259-265, 1987). The other endopeptidase, LEP-I, was purified to homogeneity by conventional chromatographic techniques from Streptococcus cremoris H61. The enzyme appeared to be monomeric, with an apparent molecular weight of 98,000, and its isoelectric point was 5.1. For the hydrolysis of αs1-CN(f1-23), the enzyme had an optimum pH and temperature of 7.0 to 7.5 and 40°C, respectively. Its activity was inhibited by such chelating agents as EDTA and 1,10-phenanthrolin, and it could be fully reactivated by Mn²⁺. Inhibitors specific for serine and thiol proteases had no effect on the protease activity. The enzyme showed a high affinity toward the Glu-Asn peptide bond of αs1-CN(f1-23) and αs1-CN(f91-100) but showed no hydrolysis activity toward αs1-CN(f1-52), αs1-CN(61-122), αs1-CN(136-196), αs1-casein, β-casein, κ-casein, α-lactalbumin, and β-lactoglobulin. The K_m and V_max of LEP-I for αs1-CN(f1-23) were 14.2 pM and 139 U, respectively.     

Purification and characterization of a novel metalloendopeptidase from Streptococcus cremoris H61. A metalloendopeptidase that recognizes the size of its substrate

An endopeptidase (LEP-II), which has a unique substrate specificity, was purified to homogeneity by conventional chromatographic techniques from Streptococcus cremoris H61. The enzyme was a metalloendopeptidase since it was inhibited by EDTA and 1,10-phenanthroline; the metal-depleted enzyme could be fully reactivated by micromolar levels of Zn2+ and was not inhibited by specific inhibitors for serine or thiol protease. The molecular mass of the enzyme was estimated to be 80 kDa by Sephacryl S-300 gel filtration and high-performance liquid chromatography with a TSK-G3000SW column. The enzyme consisted of two identical subunits and the N-terminal sequence of LEP-II was determined up to the 19th residue. Although the enzyme had a broad substrate specificity it specifically hydrolyzed the peptide bonds involving the amino groups of hydrophobic amino acid residues. Various small polypeptides, such as alpha s1-CN(f1-23), alpha s1-CN(f91-100), oxidized insulin B chain, glucagon and some biologically active peptides were hydrolyzed. However, a variety of larger polypeptides or proteins, such as alpha s1-CN(f1-54), alpha s1-CN(f61-123), alpha s1-CN(f136-196), alpha s1-casein, beta-casein, and kappa-casein were not hydrolyzed. LEP-II recognized the size of its substrates, which were limited below a molecular mass of about 3.5 kDa.     

中国七星瓢虫群聚和迁飞的数学分析

自1976年首次在秦皇岛海岸发现七星瓤虫的群聚现象以来,又经过连续8年的广泛研究,进一步讨论了七星瓤虫在黄、渤海域的群聚范围、群聚数量、群聚规律以及迁飞等问题。本文的重点是试图从数学上论证七星瓤虫的迁飞,并进一步预测预报七星瓤虫的群聚日期,以便为群聚七星瓢虫的利用奠定基础。 根据可能有关的因子设64个自变量,2个因变量,通过反复计算,大量筛选可能与虫情有关的气象因子,结果入选的气象因子说明,群聚瓢虫是迁飞过程中迫降入海,并由风浪推上海岸的。经过用双层筛选逐步回归和逐步判别两种数学方法去筛选自变量,结果基本上是一致的。因此从定量角度对七星瓢虫的群聚和迁飞给出了论证。并说明七星瓢虫的群聚与近期气象条件密切相关,可以利用短期或中期气象预报代入判别式进行群聚的预测预报。     

Ion channel expression by white matter glia: the type-1 astrocyte.

We describe the electrophysiological properties of acutely isolated type-1 astrocytes using a new "tissue print" dissociation procedure. Because the enzymes used did not destroy or modify the ion channels, and the cells retained many processes, the properties may reflect those in vivo. The types of ion channels in type-1 astrocytes changed rapidly during the first 10 postnatal days, when they attained their adult phenotype. This change was dependent on the presence of neurons. In culture, most of these channel types were not expressed, but a phenotype more typical of that in vivo could be induced by co-culture with neurons. The electrophysiological properties of astrocytes make some existing hypotheses of astrocyte function less likely.     

国产磨芋属的染色体核型报道(1)

本文报道了磨芋属(Amorphophallus Blume)六个种的染色体数目和核型,其中5个种属于首次报道。其核型公式如下: 1.滇磨芋 K(2n)=2x=26=26m.2.磨芋 K(2n)=2x=26=26m.3.攸落磨芋K(2n)=2x=26=22m(2SAT)+4sm.(2SAT).4.西盟磨芋 K(2n)=2x=26=20m+4sm+2st.5.勐海磨芋 K(2n)=2x=26=22m+4sm.6.白磨芋 K(2n)=2x=26=20m(2SAT)+6sm。     

Early-blocked sporulation mutations alter expression of enzymes under carbon control in Bacillus subtilis

Summary The physiological roles of the gene subset defined by early-blocked sporulation mutations (spo0) and their second-site suppressor alleles (rvtA11 and crsA47) remain cryptic for both vegetative and sporulating Bacillus subtilis cells. To test the hypothesis that spo0 gene products affect global regulation, we assayed the levels of carbon- and nitrogen-sensitive enzymes in wild-type and spo0 strains grown in a defined minimal medium containing various carbon and nitrogen sources. All the spo0 mutations (except spo0J) affected both histidase and arabinose isomerase levels in an unexpected way: levels of both carbon-sensitive enzymes were two- to six-fold higher in spo0 strains compared to wild type, when cells were grown on the derepressing carbon sources arabinose or maltose. There was no difference in enzyme levels with glucose-grown cells, nor was there a significant difference in levels of the carbonindependent enzymes glutamine synthetase and glucose-6-phosphate dehydrogenase. This effect was not due to a slower growth rate for the spo0 mutants on the poor carbon and nitrogen sources used. The levels of carbon-sensitive enzymes were not simply correlated with sporulation ability in genetically suppressed spo0 mutants, but the rvtA and crsA suppressors each had such marked effects on wild-type growth and enzyme levels that these results were difficult to interpret. We conclude that directly or indirectly the spo0 mutations, although blocking the sporulation process, increase levels of carbon-sensitive enzymes, possibly at the level of gene expression.     

中国人线粒体DNA的八种限制酶图及其电镜结构

尽管Anderson等人(1981)已测定了人mtDNA的全部序列,但还不能全面地反映整个人类mtDNA核苷酸序列的情况。因此,在具有代表特征的中国人mtDNA序列被测定之前,为了开展对中国人mtDNA的遗传学研究,笔者构建了中国人mtDNA的8种限制酶图。并通过电镜技术对mtDNA进行了研究,发现了一种周长为2μm的小环状DNA,推测它可能在核DNA和mtDNA之间的信息传递或衰老发生中起到某些作用。     

射击过程中主要生理特征的研究

八名射击运动员(手枪2人,步枪6人),在实弹射击训练中,连续同步地记录了脑电、心电、心率、阻抗呼吸波。同时记录了受试者持枪稳定的程度。 分析表明,射击环数是五项独立无关的指标综合作用的结果。本文首次对射击时生理指标和射击环数进行系统辨识建模分析,求出每个人的系统参数_i值。找出了每人射击时最佳生理参数,为指导射击练习和反馈训练,提供了客观生理依据。 本文设计的激光光点录相方法,可反映持枪稳定的程度,简单、方便,可在射击训练中采用。     

St14/TaqⅠRFLPs在甲型血友病基因探测与产前基因诊断中的应用

<正> 甲型血友病是最常见的遗传性凝血疾病,是由于凝血因子Ⅷ(FVⅢ)基因缺陷所致。目前对此病尚无满意的治疗方法。利用DNA分析技术进行甲型血友病基因检测与产前基因诊断对开展优生优育有重要的实际应用价值。由于FⅧ基因组织结构庞大,分子病理改变复杂,目前多采用DNA限制酶片段长度多态性(RFLPs)为遗传标志对有缺陷的FⅧ基因进行连锁分析。St 14(DXS52)是人X染色体长臂远端的一段基因外DNA序列,与FⅧ基因紧密连锁。国外已将St14/Taq Ⅰ RFLPs用于甲型血友病基因携带者的检测,但尚未见到用于产前基因诊断的报道。我们利用引进的St14片段为探针,采用简化的低脂奶粉杂交体系和DNA凝胶原     

光敏亲和标记试剂8-N_3-5′-ATP的合成

<正> 近年来,光敏亲和标记技术已被广泛应用于激素及共受体,蛋白和核酸的相互作用、酶的结构与功能、膜蛋白结构、tRNA同共合成酶的识别等研究上,鉴于ATP是某些酶的底物,因此合成光敏的ATP底物类似物将有助于国内对达些酶的结构与功能进行深入研究,为此我们合成了8-N_8-5′-ATP,现将实验结果报道如下: