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zjut-mmiao铜虫 (小有名气)
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时间紧英语翻译求助
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| 由于星期天要交开题报告了，时间特别紧，我还有很多没翻译 所以求助哪位大虾帮我翻译两页。谢谢 |
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2楼2010-03-19 16:02:50
zjut-mmiao
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3.2. Effect of solid buffer on the activity of papain PREPs The propanol rinsed enzyme preparation (PREP) procedure was found to give papain catalysts showing good activity (59.2 nmol min−1 mg−1 enzyme) in tert-butanol, even at a water activity of 0.2. The reaction rates of papain PREPs in the presence of solid buffers were plotted against their aqueous pKa values, as shown in Fig. 2. This indicates that there is no exact relation between the rate and aqueous pKa. None of the tested solid buffers increased the reaction rate of papain PREPs compared to the control reaction, although Glu/Glu-Na, PIPES/PIPES-Na and NaH2PO4/Na2HPO4 gave values close to this. Clearly low catalytic activity of the enzyme was observed in the presence of buffers that are strongly basic (pKa > 9; AMPSO/AMPSO-Na, Gly/Gly-Na and CAPSO/CAPSO-Na) or strongly acid (pKa < 4; citric acid and monosodium citrate). Interestingly, some buffers (Maleate-Na/Maleate-Na2 and TES/TES-Na) having intermediate pKa values also gave low reaction rates. Note that high catalytic activity of papain in aqueous solution is found in the pH range 4–8.5, with the limits set by the side chain pKa values of Cys-25 and His-159 [25,26]. In the optimal pH range, these active site residues exist predominantly as a thiolate-imidazolium ion pair (Cys-S−/His-ImH+). However, it should be remembered that the aqueous pKa of solid buffers is not the only factor affecting the catalytic activity. The potential for exchange of H+ and Na+ will also depend on the nature of the crystalline solid, and the solubility of the solid buffers may also be important. Fig. 3 shows the relationship between the synthetic rate of papain PREPs and the fraction of deprotonated indicator after equilibration with the buffer used. Surprisingly, the correlation between the indicator response and the catalytic rate is only slightly better than for aqueous pKa (Fig. 2). It is clear that the most basic buffers (mainly deprotonated indicator) give lower initial rates. At 0% deprotonated fraction, the reaction rates in the presence of glutamic/glutamate-Na and citric/ monosodium citrate were remarkably different. Probably the indicator is not acidic enough to sense the difference in these two buffer pairs, which is reflected in the lower aqueous pKa1 of citric acid. These results differ in two important ways from those found with subtilisin PREPs [12]. Firstly, the relationship with the potential for exchange of H+ and Na+ ions is much less clear. This indicated that the catalytic activity of papain PREPs is not so clearly determined by the solid-state buffers. Secondly, the rate falls for both the more acidic and more basic buffer pairs, whereas subtilisin activity increased steadily with increasing indicator deprotonated fraction. This was attributed to the increase in catalytic activity of subtilisin as the active site triad is deprotonated. The higher activity of papain with intermediate buffers may be because formation of the required thiolate-imidazolium ion pair at the active site is favoured. Too acidic buffers will cause protonation of Cys-25, while too basic ones will deprotonate His-159. A comment is appropriate on the consequences of the mixture of proteins present in the preparation used. To a first approximation, each might be expected to equilibrate separately with the buffers present. Thus the impurities would not have a great effect on the behaviour of a given protein. However, in the experiments here another factor might be a contribution to the measured reaction rates from other proteases present. 3.3. Effect of added acid on the rate of enzyme reaction In the reported reactions with subtilisin, an acid byproduct is produced during transesterification, and the addition of solid buffer pairs was able to limit the effects of this on the reaction [11]. The model reaction used here with papain was peptide synthesis in which an acid (Z-Gly) is consumed rather than produced. The rate was found to be reduced by spiking with acetic acid during the reaction (Fig. 4a). However, this effect was not overcome by the presence of PIPES/PIPES-Na+ as solid-state buffer, which did not change the trend of reaction compared to the control. Fig. 4b gives a clear indication that the solid-state buffer failed to control the protonation state of papain PREPs, although increasing acetic acid concentrations inhibit the reaction. The two other solid buffers giving high reaction rates, glutamic/glutamate-Na and NaH2PO4/Na2HPO4, also had no effect in counteracting the effects of added acid. The effects of acid are lower than those described for subtilisin, |
3楼2010-03-19 16:09:20
zjut-mmiao
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4楼2010-03-19 16:10:05
zjut-mmiao(金币+80):辛苦了 谢谢 2010-03-20 17:49
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3.2. 固体缓冲剂对木瓜蛋白酶活性的影响 用丙醇冲洗酶制剂（PrEP）的步骤，发现可以使木瓜蛋白酶在丁醇中显示良好的催化 活性(59.2 nmol min - 1 mg - 1 enzyme) ，即使在水中也显示了 0.2的活性。在固体缓冲剂 体系中，把木瓜蛋白酶PREPs的反应速率按溶液的 pKa值做图，如图2所示。图示表明 ，反应速率与溶液的pKa之间没有确切关系。 与控制反应相比，被测试的固体缓冲液 都不能提高papain PREPs 的反应速率，虽然 Glu/Glu-Na, PIPES/PIPES-Na 以及 NaH2PO4/Na2HPO4 的值接近了这一点。显然，可观察到此酶在强碱性(pKa > 9; AMPSO/AMPSO-Na, Gly/Gly-Na and CAPSO/CAPSO-Na) 或强酸性 (pKa < 4; citric acid and monosodium citrate) 缓冲溶液体系下的催化活性低。有趣的是，一些缓冲体系 如(Maleate-Na/Maleate-Na2 and TES/TES-Na)具有中等 pKa值也得到了较低反应速率。 值得注意的是，在pH范围4-8.5的溶液中， 发现木瓜蛋白酶具有较高的催化活性，这 主要是受侧链Cys-25 and His-159 pKa值的限制[25,26]。在最佳 pH值范围内，这些活性 位残留物主要以硫醇盐-咪唑离子对（Cys-S-/His-ImH +）形式存在 。 然而，应该记住 的是固体缓冲剂的pKa值不是唯一影响催化活性的因素。H +和Na +例子交换的可能性 也取决于晶体固体的性质，以及固体缓冲剂的溶解性可能也很重要。图 3 显示了当用 缓冲液平衡后，木瓜papain PREPs 合成率与去质子指示剂指示分数之间的关系。出人 意外的是，指标剂响应与催化率之间的相关性只是略优于溶液的pKa值（图2）。显然 ， 最基本的缓冲剂（主要是去质子指标剂）提供了较低的初始速率。当去质子分数 为0％时，glutamic/glutamate-Na and citric/monosodium citrate 缓冲对的反应率显著不同 。也许是由于指标剂不够酸性去检测到这两个缓冲对的差异，这也反映在具有较低的 溶液pKa1的柠檬酸体系中。 这些结果与在subtilisin PREPs体系中的发现存在两个不同的重要途径 [12]。首先，H + 和Na +离子可能交换的关系不太清楚。这表明，papain PREPs 的催化活性不是很明显 地取决于固态缓冲剂。 其次，papain PREPs 的催化活性在更酸和更碱缓冲对体系下反 应速率下降，而subtilisin 蛋白酶的活性随指标去质子分数的增长稳步提高。这是由于 随着活性位triad去质子化，subtilisin的催化活性提高。在中等缓冲剂下，papain 有较高 的活性，可能是因为在中级缓冲体系下有利于在活性位形成了所需的thiolate- imidazolium离子。过酸的缓冲剂将引起Cys-25的质子化, 同时过碱的缓冲剂将使 His-159 脱质子。 应该适当的评价一下在合成中所使用蛋白质的混合所引起的后果。大致上，若每一个 蛋白质都如预期的那样分别平衡存在缓冲液中。此时，杂质将不会对某一 蛋白质行 为有很大影响。然而，在此实验中的另一个因素可能是其他蛋白酶存在对测量反应速 率做出贡献。 3.3. 所加入的酸对酶反应速率的影响 在所报道的subtilisin反应中，在酯交换过程中产生了酸副产品，而固体缓冲离子对的 添加可以限制副反应的发生 [11]. 在这里使用的模型反应，papain 与多肽合成，其中一 酸 (Z-Gly) 是消耗而不是生产。发现在反应过程中，反应速率通过滞留乙酸而降低(Fig. 4a). 然而，这种效应并没有通过添加However, this effect was not overcome by the PIPES/PIPES-Na+ 作为固态缓冲剂而解决, 它们并没有改变反应趋势较对照实验。Fig. 4b 给出了明确的迹象表明，固态缓冲剂未能控制papain PREPs的质子化状态，虽然增 加醋酸浓度抑制反应。另外两种固体缓冲剂得到了高的反应速率, glutamic/glutamate- Na 和 NaH2PO4/Na2HPO4, 同时对添加酸的效应没有有任何抑制影响作用。酸的影响 也低于在subtilisin体系下所描述的， --------------------------------------------- ---------------------------------------------- 注：非该领域人士翻译，难免有不准确之处，请楼主自行更正。  |
5楼2010-03-19 20:31:16