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31.
Purification and Characterization of a Substrate-Size-Recognizing Metalloendopeptidase from Streptococcus cremoris H61 总被引:6,自引:5,他引:1 下载免费PDF全文
Tsong-Rong Yan Norihiro Azuma Shuichi Kaminogawa Kunio Yamauchi 《Applied microbiology》1987,53(10):2296-2302
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 Km and Vmax of LEP-I for αs1-CN(f1-23) were 14.2 pM and 139 U, respectively. 相似文献
32.
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. 相似文献
33.
34.
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。 相似文献
35.
<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 相似文献
36.
<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: 相似文献
37.
测定了3T3细胞、人和大鼠一些组织中DNA拓扑异构酶Ⅰ的活性;估计了核酸内切酶对拓扑酶Ⅰ松弛活性测定的干扰程度;发现增殖组织全细胞抽提液中酶比活高于正常分化组织,而且在异常增殖组织中酶比活的增高更为显著。 相似文献
38.
An X-prolyl dipeptidyl aminopeptidase (X-PDAP; EC 3.4.14.5) was identified to be loosely bound on the inner cell membrane fraction of Lactococcus lactis subsp. cremoris nTR. The biosynthesis of X-PDAP was continuously increased before the late-log growth phase of the bacteria. Both Gly-Pro-pNA and Ala-Ala-pNA were hydrolyzed by X-PDAP; the kcat/Km value of the former was about 10-fold that of the latter. The Ki of X-Pro and Pro-X were more specific to X-PDAP than those of X-Ala. The enzyme splitting a dipeptide sequentially from beta-casomorphin as a model catalytic pattern was identified and some properties of the enzyme were further characterized. 相似文献
39.
Renmei Ren Kanhui Yan Yanjie Su Hanjun Qi Bin Liang Wunyong Bao Frans B. M. de Waal 《Primates; journal of primatology》1991,32(3):321-327
Observations were made following 130 spontaneous aggressive incidents in two small breeding groups of captive golden monkeys
(Rhinopithecus roxellanae roxellanae). Participants were observed both during the first 10 min following these incidents and during matched control observations.
An increased contact rate was observed between opponents following the aggressive incident. Post-conflict contacts were characterized
by a number of behavior patterns: open mouth, rapid grooming, embrace, and crouching. Adult males played an important role
as mediator in agonistic disputes among females: males intervened in 93.6% of female fights. It is speculated that this intervention
behavior is related to the species' organization into one-male units. 相似文献
40.
Yan Ming Ling Yihling Huang Shansheng Gong Shuqing Xu Zhenxing 《Biological trace element research》1991,29(3):281-288
Contents of Cu and Zn of into-pulmonary blood (IPB), out-pulmonary blood (OPB), Lung tissue, and supernatant and macrophages of Lung Lavage were determined in superior mesenteric artery occlusion (SMAO) shock of rabbits. Zn of pulmonary tissue was 11.42 +/- 0.60 and 14.52 +/- 1.78 (micrograms/g wet wt) in SMAO shock and control groups, respectively. Content of Zn was found to be lower, Cu was not changed, and Cu/Zn ratio increased in lung tissue in SMAO shock. Contents of Cu and Zn in other samples were not changed. The results suggest that lower Zn in lung tissue related to acute lung injury. 相似文献