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Wu X Kobori H Orita I Zhang C Imanaka T Xing XH Fukui T 《Biotechnology and bioengineering》2012,109(1):53-62
A novel thermostable NAD(P)H oxidase from the hyperthermophilic archaeon Thermococcus kodakarensis KOD1 (TkNOX) catalyzes oxidation of NADH and NADPH with oxygen from atmospheric air as an electron acceptor. Although the optimal temperature of TkNOX is >90°C, it also shows activity at 30°C. This enzyme was used for the regeneration of both NADP(+) and NAD(+) in alcohol dehydrogenase (ADH)-catalyzed enantioselective oxidation of racemic 1-phenylethanol. NADP(+) regeneration at 30°C was performed by TkNOX coupled with (R)-specific ADH from Lactobacillus kefir, resulting in successful acquisition of optically pure (S)-1-phenylethanol. The use of TkNOX with moderately thermostable (S)-specific ADH from Rhodococcus erythropolis enabled us to operate the enantioselective bioconversion accompanying NAD(+) regeneration at high temperatures. Optically pure (R)-1-phenylethanol was successfully obtained by this system after a shorter reaction time at 45-60°C than that at 30°C, demonstrating an advantage of the combination of thermostable enzymes. The ability of TkNOX to oxidize both NADH and NADPH with remarkable thermostability renders this enzyme a versatile tool for regeneration of the oxidized nicotinamide cofactors without the need for extra substrates other than dissolved oxygen from air. 相似文献
114.
Evidence of functional selection pressure for alternative splicingevents that accelerate evolution of protein subsequences 下载免费PDF全文
Recently, it was proposed that alternative splicing may act as a mechanism for opening accelerated paths of evolution, by
reducing negative selection pressure, but there has been little evidence so far whether this could produce adaptive benefit.
Here we employ metrics of very different types of selection pressures (e.g. against amino acid mutations (Ka/Ks); against mutations at synonymous sites (Ks); and for protein reading-frame preservation) to address this question via genome-wide analyses of human, chimpanzee, mouse,
and rat. These data show that alternative splicing relaxes Ka/Ks selection pressure up to seven-fold, but intriguingly that this effect is accompanied by a strong increase in selection pressure against synonymous mutations, which propagates into the adjacent intron, and correlates strongly with
the alternative splicing level observed for each exon. These effects are highly local to the alternatively spliced exon. Comparisons
of these four genomes consistently show an increase in the density of amino acid mutations (Ka) in alternatively spliced exons, and a decrease in the density of synonymous mutations (Ks). This selection pressure against synonymous mutations in alternatively spliced exons was accompanied in all four genomes
by a striking increase in selection pressure for protein reading-frame preservation, and both increased markedly with increasing
evolutionary age. Restricting our analysis to a subset of exons with strong evidence for biologically functional alternative
splicing produced identical results. Thus alternative splicing apparently can create evolutionary “hotspots” within a protein
sequence, and these events have evidently been selected for during mammalian evolution. 相似文献
115.
大量研究表明,TGF-β作为一种具有多种功能的生长因子,是全身瘢痕愈合的重要刺激因素.在人眼部参与多种眼部纤维化的过程。尤其在促进青光眼滤过术后结膜瘢痕形成中起重要作用。Smad蛋白家族在TGF-β超家族的信号转导中具有重要的作用。Smad7主要通过抑制TGF书受体介导的Smad2、Smad3磷酸化来拮抗TGF-β的信号转导,被认为是TGF-β超家族信号转导自我调节的一种负反馈信号。 相似文献
116.
Ning Xu Yijie Dong Xinxin Cheng Qilin Yu Kefan Qian Jiwei Mao Chang Jia Xiaohui Ding Bing Zhang Yulu Chen Biao Zhang Laijun Xing Mingchun Li 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2014
Iron bioavailability is crucial for mitochondrial metabolism and biosynthesis. Dysregulation of cellular iron homeostasis affects multiple aspects of mitochondrial physiology and cellular processes. However, the intracellular iron trafficking pathway in Candida albicans remains unclear. In this study, we characterized the Mrs4–Ccc1–Smf3 pathway, and demonstrated its important role in maintaining cellular iron levels. Double deletion of vacuolar iron exporter SMF3 and mitochondrial iron transporter MRS4 further elevated cellular iron levels in comparison with the single MRS4 deletion. However, deletion of vacuolar iron importer CCC1 in the mrs4?/? mutant restored cellular iron homeostasis to normal wild-type levels, and also normalized most of the defective phenotypes in response to various environmental stresses. Our results also suggested that both Mrs4 and Ccc1 contributed to the maintenance of mitochondrial function. The mrs4?/? and mrs4?/?smf3?/? mutants exhibited an obvious decrease in aconitase activities and mitochondrial membrane potential, whereas deletion of CCC1 in the mrs4?/? mutant effectively rescued these defects. Furthermore, we also found that the Mrs4–Ccc1–Smf3 pathway was indispensable for cell-wall stability, antifungal drug tolerance, filamentous growth and virulence, supporting the novel viewpoint that mitochondria might be the promising target for better antifungal therapies. Interestingly, the addition of exogenous iron failed to rescue the defects on non-fermentable carbon sources or hyphae-inducing medium, indicating that the defects in mitochondrial respiration and filamentous development might result from the disturbance of cellular iron homeostasis rather than environmental iron deprivation. Taken together, our results propose the Mrs4–Ccc1–Smf3 pathway as a potentially attractive target for antifungal drug development. 相似文献
117.
Qiaoli Feng ;Zi Fang ;Zhenguang Yan ;Rui Xing ;Liping Xie ;Rongqing Zhang 《Acta biochimica et biophysica Sinica》2009,(11):955-962
We previously identified a matrix protein, MSI7, from pearl oyster Pinctada fucata. According to the structural analysis, the DGD site in the N-terminal of MSI7 is crucial for its role in the shell formation. In this study, we expressed a series of recombinant MSI7 proteins, including the wild-type and several mutants directed at the DGD site, using an Escherichia coli expression system to reveal the structure-function relationship of MSI7. Furthermore, in vitro crystallization, crystallization speed assay, and circular dichroism spectrometry were carried out. Results indicated that wild-type MSI7 could induce the nucleation of aragonite and inhibit the crystallization of calcite. However, none of the mutants could induce the nucleation of aragonite, but all of them could inhibit the crystallization of calcite to some extent. And all the proteins accelerated the crystallization process. Taken together, the results indicated that MSI7 could contribute to aragonite crystallization by inducing the nucleation of aragonite and inhibiting the crystallization of calcite, which agrees with our prediction about its role in the nacreous layer formation of the shell. The DGD site was critical for the induction of the nucleation of aragonite. 相似文献
118.
HDL carries biologically active lipids such as sphingosine-1-phosphate (S1P) and stimulates a variety of cell signaling pathways in diverse cell types, which may contribute to its ability to protect against atherosclerosis. HDL and sphingosine-1-phosphate receptor agonists, FTY720 and SEW2871 triggered macrophage migration. HDL-, but not FTY720-stimulated migration was inhibited by an antibody against the HDL receptor, SR-BI, and an inhibitor of SR-BI mediated lipid transfer. HDL and FTY720-stimulated migration was also inhibited in macrophages lacking either SR-BI or PDZK1, an adaptor protein that binds to SR-BI''s C-terminal cytoplasmic tail. Migration in response to HDL and S1P receptor agonists was inhibited by treatment of macrophages with sphingosine-1-phosphate receptor type 1 (S1PR1) antagonists and by pertussis toxin. S1PR1 activates signaling pathways including PI3K-Akt, PKC, p38 MAPK, ERK1/2 and Rho kinases. Using selective inhibitors or macrophages from gene targeted mice, we demonstrated the involvement of each of these pathways in HDL-dependent macrophage migration. These data suggest that HDL stimulates the migration of macrophages in a manner that requires the activities of the HDL receptor SR-BI as well as S1PR1 activity. 相似文献
119.
两种沼虾溶菌酶基因ORF的克隆和罗氏沼虾溶菌酶基因的组织表达(英文) 总被引:8,自引:0,他引:8
分别提取罗氏沼虾和日本沼虾血细胞总RNA,RT-PCR扩增获得特异性cDNA片段,纯化后克隆到T载体上。序列测定表明所克隆的两种沼虾溶菌酶基因的开放阅读框(ORF)为477bp,共编码158个氨基酸,包括溶菌酶成熟肽140个氨基酸残基和信号肽18个氨基酸残基。同源性分析表明,罗氏沼虾和日本沼虾溶菌酶基因的碱基序列及推测氨基酸序列高度同源,分别为99.4%和98.1%。两种沼虾溶菌酶基因的碱基序列和推测氨基酸序列与Gen-Bank上其他对虾溶菌酶的同源性达83.0%和80.0%以上。两种沼虾溶菌酶都具有c-型溶菌酶典型的两个酶活性位点(Glu51)和(Asp68),以及8个保守结构氨基酸残基Cys,且在101、106和107位上缺少Asp,因而推测本实验所克隆的两种沼虾溶菌酶基因属c-型溶菌酶基因的非钙结合亚型。以PCR法制备罗氏沼虾溶菌酶基因的生物素标记探针,斑点杂交检测感染弧菌后溶菌酶基因mRNA在各组织中的转录水平,结果表明受感染6h后在眼、肌肉、鳃、肝胰腺、肠管中的表达量均有升高,其中在肝胰腺中的表达量最高,约为对照组的560%。在不同感染时间里,肝胰腺中该基因表达量有较大的变化:感染后3h表达量最低,24h后表达量升至最高,大约为对照组的430%,48h时的表达量又有所下降,但仍明显高于对照组(约为330%)。受弧菌感染后罗氏沼虾溶菌酶基因转录的上调证明溶菌酶基因在非特异性免疫中的直接作用,同时表明肝胰腺可能在沼虾的免疫防御过程起重要作用。
相似文献
120.
Kong X Zhang C Jin X Wu X Zhang S Zhong Z Feng Q Liu T Yuan H 《BioFactors (Oxford, England)》2011,37(4):323-327
The objective of this study is to observe the effect of high-mobility group protein B1 A Box (HMGB1 A) box on lung injury in mice with acute pancreatitis and its effect on the level of high-mobility group protein B1 (HMGB1) in lung, to explore the mechanism. A total of 60 male Institute of Cancer Research mice were randomly divided into control group (n = 30) and treatment group (n = 30). Severe acute pancreatitis mice model was induced by 20% L-Arg intraperitoneal injection. The recombination HMGB1 A box was used in treatment after modeling. All the mice were killed under anesthesia at 24 and 48 h after the modeling injection. The level of HMGB1 and activity of myeloperoxidase (MPO) in lung were measured. The pathological changes of lung were observed. The level of HMGB1 in lung of A box treatment group decreased more significantly 24 h and 48 h after modeling compared with control group. The activity of MPO in lung of A box treatment group decreased more significantly 24 h after modeling compared with control group. The lung tissue pathologic score of A box treatment group decreased more significantly 48 h after modeling compared with control group. HMGB1 expression levels in the lungs were positively related to histological score of injured lung in acute pancreatitis. It indicates that HMGB1 A box is remarkably protective to lung injury induced by acute pancreatitis. 相似文献