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| 1 | Critical Amino Acid Residues for Nicotine 5'-Hydroxylation in Human CYP2A Enzymes显示文摘Objective: We have continued previous work in which we demonstrated that #117 and #372 amino acids contrib- uted to the high activities of human CYP2A13 in catalyzing 4-methylnitrosamino-1-(3-pyridyl)-1-butanone(NNK) and aflatoxin B1(AFB1) carcinogenic activation. The present study was designed to identify other potential amino acid residues that contribute to the different catalytic characteristics of two CYP2A enzymes, CYP2A6 and CYP2A13, in nicotine metabolism and provide insights of the substrate and related amino acid residues interactions. Methods: A series of reciprocally substituted mutants of CYP2A6Ile300 → Phe, CYP2A6Gly301Ala, CYP2A6Ser369 → Gly, CYP2A13Phe300 → Ile, CYP2A13Ala301 → Gly and CYP2A13Gly369 → Ser were generated by site-directed mutagenesis/baculovirus-Sf9 insect cells expression. Comparative kinetic analysis of nicotine 5'hydroxylatin by wild type and mutant CYP2A proteins was performed. Results:All amino acid residue substitutions at 300, 301 and 369 caused significant kinetic property changes in nicotine metabolism. While CYP2A6Ile300 → Phe and CYP2A6Gly301→Ala mutations had notable catalytic efficiency increases compared to that for the wild type CYP2A6, CYP2A13Phe300→Ile and CYP2A13Ala301→Gly replacement introduced remarkable catalytic efficiency decreases. CteIunfYtf iaioPcdni2de Aiontif1co #3yn3G ,v 6ali9ylal 3 r6tae9h s→seiisdge uSn ecei afrsit.iac gClanyonitnfti cicKc leaumfns fdtiilcoeyinc ea:rn#fecf3aey0sc 0eate ,l vtd#ee 3rbra0sot1uitoh san KnVsd mmw a#axe 3nree6dn 9 chV aarmuenassxcei evddmau bleueysen iVstn,. m whaCxu hYvmialPaer2in aAt htCi6eoYS noePspr 2r3pa6A9oth→6sei/t1reG 3t h lepyafl fnaien yKcc tirmsme cawphsoeea rrnttegha eenss etc. e raSontual ewlbyssi tttiiinhc- nicotine 5'-oxidation. Switching #300 or #301 residues did not affect the CYP2A protein affinities toward nicotine, although these amino acids are located in the active center. Ser369 to Gly substitution indirectly affected nicotine binding by creating more space and conformational flexibility for the nearby residues, such as Leu370 which is crucial for many hydroxylations. | Xiaoyang Hea Xu Xu Jian Shen Li Sun Anthony Y. H. Lu Clifford Weisel Junyan Hong | 2008 | Journal of Nanjing Medical University2008，22，6： | 1 |
| 2 | Mitochondrial 12S rRNA variants in 1642 Han Chinese pediatric subjects with aminoglycoside-induced and nonsyndromic hearing loss显示文摘 | Jianxin Lu Zhiyuan Li Yi Zhu Aifen Yang Ronghua Li Jing Zheng Qin Cai Guanghua Peng Wuwei Zheng Xiaowen Tang Bobei Chen Jianfu Chen Zhisu Liao Li Yang Yongyan Li Junyan You Yu Ding Hong Yu Jindan Wang Dongmei Sun Jianyue Zhao Ling Xue Jiying Wang Min-Xin | 2010 | Mitochondrion2010，，4： | 1 |
| 3 | Cytotoxicity of acrylamide and its epoxide glycidamide in CHO cells expressing human cytochrome P450 2E1显示文摘客观:调查 CYP2E1 是否 Flp 在里面为 acrylamidemetabolic 激活在负责 CHO 房间系统。方法:CYP2E1 cDNA 是从人的肝克隆的潜水艇全身的 cDNA 图书馆并且随后 transfected 进 Flp 在里面产生 CYP2E1 的稳定的 transfectant 的 CHO 房间。CYP2E1 mRNA 表示被 RT-PCR 决定。它的 epoxide glycidamide 导致了的 Acrylamideand 在 G2/M 的 cytotoxicity 和房间周期拘捕被进行分别地使用山试金和流动 cytometry。结果;在稳定地表示 CYP2E1 (CHO-2E1 ) 的 CHO 房间,当时,乐队的一种~ 1 .5 kb 尺寸在房间从 mRNA 被检测在 CHO 向量房间的没有相应乐队,它显示 CYP2E1 是成功地在 CHOcells 的 transfected。与 CHO 向量房间相比,压克力酰胺在 CHO-2E1 房间显示出生存能力的集中依赖者损失,但是 G2/M 拘捕的重要变化都没被发现。是期望, glycidamide 房间在两个导致了 cytotoxicity 的类似的侧面,并且 G2/M 拘捕介绍了在 CHO-2E1 房间增加的一个集中依赖者。结论:结果建议 thatCYP2E1 可能为压克力酰胺新陈代谢负责,并且它的代谢物 glycidamide 是一直接细胞毒素并且 genotoxic 代理人。acrylamide-inducedtox-icity 是否面对 CYP2E1 通过它的 epoxide glycidamide,应该进一步被考虑。 | Shoulin Wang Xiaoyang He Xinru Wang Junyan Hong | 2006 | Journal of Nanjing Medical University2006，20，6： | 1 |
| 4 | Influences of V5-epitope tag on the metabolic activation of AFB1 by human cytochrome P450 2A13显示文摘Objective: To explore the impact of V5-epitope tag inserted in the commercial pcDNA5/FRT/V5-His TOPO expression vector on the metabolic activation of AFB1 by human CYP2A13. Methods: A C-terminal 6 × Histag was first introduced into CYP2A13 cDNA by PCR and subsequently transferred into the expressing vector pcDNA5/FRT. Another commercial pcDNA5/FRT/V5-His TOPO expression vector was used to develop the construct directly via PCR. Both of the constructs were then transfected into Flp-In CHO and allowed for the stable expression of CYP2A13. The mouse CYP2A5 and the vector alone were used as positive and negative control, respectively. The presence of CYP2A5 and CYP2A13 cDNA and their protein expression in the stable transfectant cells were determined by immunoblotting assay using a monoclonal antibody against 6 × Histag. The AFB1-induced cytotoxicity in these tranfected CHO cells were conducted by MTS assay and the IC50 of cell viability was used to compare the CYP enzyme metabolic activity in AFB1 metabolism among these cells. Results: In accordance with the Flp-In system working mechanism, all the transfectant cells presented same protein expression level. The CHO cells expressing CYP2A5 was more sensitive to AFB1 treatment than those cells expressing CYP2A13, there was about 30-fold IC50 difference between the two cells(2.1 nmol/L vs 58 nmol/L). Interestingly, CYP2A13 fused with V5-Histag had the lost of metabolic activity to AFB1 than that fused with Histag alone, the IC50 of the viability in CHO-2A13-His-V5 cells was about 20-fold less than CHO-2A13-His(> 1 000 nmol/L vs 58 nmol/L). However, there was no change between CYP2A5 fused with V5-Histag and Histag alone (2.4 nmol/L vs 2.1 nmol/L). Conclusion: The results demonstrate that CYP2A13 fused with V5-epitope has a significant impact on its metabolic activation to AFB1, which indicated that it should be careful to select a new expressing vector for evaluating the enzyme activity in carcinogen metabolism. | Shoulin Wang Xiaoyang He Xinru Wang Junyan Hong | 2006 | Journal of Nanjing Medical University2006，20，5： | 0 |
| 5 | Dock4 is required for the maintenance of cochlear hair cells and hearing function显示文摘Auditory hair cells(HCs)are the mechanosensory receptors of the cochlea,and HC loss or malfunction can result from genetic defects.Dock4,a member of the Dock180-related protein superfamily,is a guanine nucleotide exchange factor for Rac1,and previous reports have shown that Dock4 mutations are associated with autism spectrum disorder,myelodysplastic syndromes,and tumorigenesis.Here,we found that Dock4 is highly expressed in the cochlear HCs of mice.However,the role of Dock4 in the inner ear has not yet been investigated.Taking advantage of the piggyBac transposon system,Dock4 knockdown(KD)mice were established to explore the role of Dock4 in the cochlea.Compared to wild-type controls,Dock4 KD mice showed significant hearing impairment from postnatal day 60.Dock4 KD mice showed hair bundle deficits and increased oxidative stress,which eventually led to HC apoptosis,late-onset HC loss,and progressive hearing loss.Furthermore,molecular mechanism studies showed that Rac1/β-catenin signaling was significantly downregulated in Dock4 KD cochleae and that this was the cause for the disorganized stereocilia and increased oxidative stress in HCs.Overall,our work demonstrates that the Dock4/Rac1/β-catenin signaling pathway plays a critical role in the maintenance of auditory HCs and hearing function. | Guodong Hong Xiaolong Fu Jieyu Qi Buwei Shao Xuan Han Yuan Fang Shuang Liu Cheng Cheng Chengwen Zhu Junyan Gao Xia Gao Jie Chen Ming Xia Wei Xiong Renjie Chai | 2023 | Fundamental Research2023，3，4： | 0 |
| 6 | Differential regulation of JAK1 expression by ETS1 associated with predisposition to primary biliary cholangitis显示文摘Primary biliary cholangitis(PBC)is an autoimmune liver disease characterized by the destruction of intrahepatic small bile ducts and progressive cholestasis,eventually leading to liver cirrhosis and hepatic failure without appropriate treatment(Terziroli Beretta-Piccoli et al.,2019). | Peng Jiang Chan Wang Mingming Zhang Ye Tian Weifeng Zhao Junyi Xin Yexi Huang Zhibin Zhao Wenjuan Sun Jie Long Ruqi Tang Fang Qiu Xingjuan Shi Yi Zhao Li Zhu Na Dai Lei Liu Xudong Wu Jinshan Nie Bo Jiang Youlin Shao Yueqiu Gao Jianjiang Yu Zhigang Hu Zhidong Zang Yuhua Gong Yaping Dai Lan Wang Ningling Ding Ping Xu Sufang Chen Lu Wang Jing Xu Luyao Zhang Junyan Hong Ruonan Qian Hu Li Xuan Jiang Congwei Chen Wenyan Tian Jian Wu Yuzhang Jiang Chongxu Han Kui Zhang Hong Qiu Li Li Hong Fan Liming Chen Jianqiong Zhang Zhongsheng Sun Xiao Han Zhenhua Dai Erguang Li M.Eric Gershwin Zhexiong Lian Xiong Ma Michael F.Seldin Weichang Chen Meilin Wang Xiangdong Liu | 2023 | Journal of Genetics and Genomics2023，50，10： | 0 |
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