榆黄蘑对小鼠血乳酸,血尿素,乳酸脱氢酶影响的实验研究

以小鼠为实验材料，研究榆黄蘑发酵液对血乳酸、乳酸脱氢酶（ＬＤＨ）、血尿素（ＢＵＮ）的影响。结果表明，与对照组比较，服用榆黄蘑发酵液２５ｄ的小鼠剧烈运动停止后５０ｍｉｎ，血乳酸含量降低３２％（Ｐ＜０．０５）；乳酸在血液中的清除速率提高１９．５％（Ｐ＜０．０１）；血清ＬＤＨ活性提高１１．６％（Ｐ＜０．０５）。ＢＵＮ增量降低６５％（Ｐ＜０．０５）。实验结果提示榆黄蘑有较好的抗疲劳作用。     

budC敲除及ldhA过表达对Klebsiella pneumoniae合成D-乳酸的影响

为了提高K. pneumoniae中D-乳酸的合成效率,本文以BUD和LDH为改造目标,扩增丁二醇脱氢酶基因budC,并在其中插入四环素抗性基因tet,构建了基因敲除载体pTBT,转化K. pneumoniae,利用同源重组技术,敲除K. pneumoniae染色体上的budC基因,得到重组菌K. pneumonia B-;在此基础上,构建了表达载体pKP-ldhA,转化K. pneumoniae B-,过量表达乳酸脱氢酶基因ldhA,得到重组菌K. pneumoniae B-L+。摇瓶发酵结果显示,重组菌K. pneumoniae B-L+的丁二醇合成浓度比原始菌降低了90%以上,D-乳酸合成浓度比K. pneumoniae B-和原始菌分别提高了77.1%和41.4%,发酵罐实验D-乳酸产量68.4g/L,转化率0.78,生产强度1.22g/(L·h)。结果表明,敲除budC及过表达ldhA有利于改善克雷伯肺炎杆菌中D-乳酸的合成。     

Pyruvate dehydrogenase kinase regulatory mechanisms and inhibition in treating diabetes, heart ischemia, and cancer

The fraction of pyruvate dehydrogenase complex (PDC) in the active form is reduced by the activities of dedicated PD kinase isozymes (PDK1, PDK2, PDK3 and PDK4). Via binding to the inner lipoyl domain (L2) of the dihydrolipoyl acetyltransferase (E2 60mer), PDK rapidly access their E2-bound PD substrate. The E2-enhanced activity of the widely distributed PDK2 is limited by dissociation of ADP from its C-terminal catalytic domain, and this is further slowed by pyruvate binding to the N-terminal regulatory (R) domain. Via the reverse of the PDC reaction, NADH and acetyl-CoA reductively acetylate lipoyl group of L2, which binds to the R domain and stimulates PDK2 activity by speeding up ADP dissociation. Activation of PDC by synthetic PDK inhibitors binding at the pyruvate or lipoyl binding sites decreased damage during heart ischemia and lowered blood glucose in insulin-resistant animals. PDC activation also triggers apoptosis in cancer cells that selectively convert glucose to lactate. Received 25 August 2006; received after revision 20 November 2006; accepted 20 December 2006     

Alcohol dehydrogenase 2 is a major hepatic enzyme for human retinol metabolism

The metabolism of all-trans- and 9-cis-retinol/ retinaldehyde has been investigated with focus on the activities of human, mouse and rat alcohol dehydrogenase 2 (ADH2), an intriguing enzyme with apparently different functions in human and rodents. Kinetic constants were determined with an HPLC method and a structural approach was implemented by in silico substrate dockings. For human ADH2, the determined K_m values ranged from 0.05 to 0.3 μM and k_cat values from 2.3 to 17.6 min⁻¹, while the catalytic efficiency for 9-cis-retinol showed the highest value for any substrate. In contrast, poor activities were detected for the rodent enzymes. A mouse ADH2 mutant (ADH2Pro47His) was studied that resembles the human ADH2 setup. This mutation increased the retinoid activity up to 100-fold. The K_m values of human ADH2 are the lowest among all known human retinol dehydrogenases, which clearly support a role in hepatic retinol oxidation at physiological concentrations. Received 12 October 2006; received after revision 6 December 2006; accepted 8 January 2007     

Diversity in enoyl-acyl carrier protein reductases

The enoyl-acyl carrier protein reductase (ENR) is the last enzyme in the fatty acid elongation cycle. Unlike most enzymes in this essential pathway, ENR displays an unusual diversity among organisms. The growing interest in ENRs is mainly due to the fact that a variety of both synthetic and natural antibacterial compounds are shown to specifically target their activity. The primary anti-tuberculosis drug, isoniazid, and the broadly used antibacterial compound, triclosan, both target this enzyme. In this review, we discuss the diversity of ENRs, and their inhibitors in the light of current research progress. Received 3 November 2008; received after revision 5 December 2008; accepted 8 December 2008     

烟酸羟基化菌株睾酮丛毛单胞菌JA1抗性质粒的消除

睾酮丛毛单胞菌JA1可羟基化烟酸产生6-羟基烟酸,并且对多种抗生素具有抗性.碱变性法提取JA1细胞内的质粒, 检测到一个约19 kb大小的质粒.用0.002 5% SDS进行质粒消除实验,结果表明,质粒消除后菌株对卡那霉素敏感,但仍对氨苄青霉素、安普霉素和链霉素具有抗性,推测质粒携带卡那霉素的抗性基因.同时,质粒消除后,JA1菌株烟酸脱氢酶酶活性没有变化,推测烟酸脱氢酶的基因并不在质粒上.     

基于细菌脱氢酶活性法的水质综合毒性快速测定仪

在无氧条件下,生物染料亚甲基蓝可以作为生物新陈代谢过程中体内脱氢酶催化反应的电子受体,亚甲基蓝与细菌悬液共存时,其退色速度标志着细菌脱氢酶活性的大小。脱氢酶的活性大小在一定程度上能够反映出微生物新代谢的能力,微生物所处的水环境中毒物的存在会引起微生物脱氢酶活性变化,毒物的毒性与脱氢酶的活性相关。本文以动力学比色法测定亚甲基蓝的退色速度为基础设计并制造了一台水质综合毒性相关。本文以动力学比色法测定亚     

A novel salicylaldehyde dehydrosenase-NahV involved in catabolism of naphthalene from Pseudomonas putida ND6

A novel salicylaldehyde dehydrogenase involved in catabolism of naphthalene from Pseudomonas putida ND6, NahV, has been identified. NahV exhibited lower identity in amino acid sequence with the classic salicylaldehyde dehydrogenase, NahF, from P. putida ND6. This is the first report of an isofunctional enzyme of bacterial salicylaldehyde dehydrogenase. Comparison of Km and Vmax values of NahV and NahF demonstrated that NahF has a more efficient catalytic reaction than NahV, while NahV has much higher affinity for salicylaldehyde and NAD^＋. Both enzymes exhibited broad substrate specificities and catalyzed the oxidation of salicylaldehyde, 5-chlorosalicylaldehyde, formaldehyde, m-nitrobenzaldehyde, o-nitrobenzaldehyde, o-methoxybenxaldehyde, glutaraldehyde, caprylic aldehyde, and glyoxal. However, the relative rates at which the substituted analogs are transformed differ considerably. NahV activity could be enhanced by Fe^2＋, Cu^2＋ and Zn^2＋; whereas NahF activity could only be stimulated by Fe^2＋, NahF is more stable than NahV at elevated temperatures. Dot-blot hybridization analyses showed that nahF-like genes occurred in all naphthalene-degradation bacteria isolated in this study, whereas nahV-like genes were present in only some naphthalene-degrading bacteria.     

盘基网柄菌(Dictyostelium discoideum)KAX-3和AK127细胞形态发生和同工酶的比较研究

比较了盘基网柄菌(Dictyostelium discoideum)野生型细胞KAx-3和突变型细胞AK127在多细胞发育过程中的形态发生.两者有明显的区别;突变细胞的发育只能停留在细胞疏松聚集阶段,而野生型细胞能顺利完成整个发育过程.利用SDS-PAGE电泳比较分析了两者在重要发育阶段的三种同工酶:醋酶(EST)、苹果酸脱氢酶(MDH)和谷氨酸脱氢酶(GDH).结果发现AK127细胞发育阶段含有两种醋酶成分a和p,而KAX-3细胞中只有一种醋酶a;而且AK127细胞中苹果酸脱氢酶(MDH)含量在发育后14 h和16 h均高于KAX-3细胞;谷氨酸脱氢酶(GDH)同工酶在发育后14 h时相对含量高于KAX-3细胞,而在发育后16 h含量却低于KAX-3细胞.这些数据显示了两种类型细胞的同工酶有明显的差异;表明突变细胞基因的表达发生了一定的改变,由此说明了gp150分子在盘基网柄菌细胞发育中有重要作用.     

外源乳酸脱氢酶基因在双歧杆菌中的表达

以大肠杆菌 双叉双歧杆菌的穿梭质粒pHJ为基础,插入从Lactococcuslactis总DNA中用PCR方法扩增得到的报告基因乳酸脱氢酶基因(ldh),构建了双叉双歧杆菌的表达质粒pHJ LDH,并采用电转化法导入双叉双歧杆菌进行了表达研究.对双叉双歧杆菌总蛋白的SDS PAGE分析结果表明,ldh报告基因的表达产物形成了明显的条带,Western blotting结果确认了该条带为ldh基因的产物.LDH的酶活性定量测定结果表明,重组双叉双歧杆菌的蛋白粗抽提液中LDH的比活性为5.5U/mg,菌中LDH的表达量约为2mg/L.此工作为今后双叉双歧杆菌的基因工程奠定了基础.