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反向点杂交法检测中国人N-乙酰化酶基因型 总被引:3,自引:3,他引:3
目的 建立快速方便的反向点杂交法 (RDB)检测中国人N 乙酰化酶 (NAT2 )基因型。方法 采用PCR法扩增生物素标记的DNA片段 ,与固定在尼龙膜上的等位基因特异性寡核苷酸探针进行杂交 ,通过非同位素法显色检测杂交结果 ,分析NAT2 5种主要突变位点。用本法对 4 8名肺结核患者NAT2基因型进行了分析。结果 RDB法与等位基因特异扩增法结果完全一致。 4 8名肺结核患者NAT2 5、 6、 73种突变型等位基因的基因频率分别为 1 0 4 %、2 2 9%和 15 6 %。野生型纯合子、杂合子和突变型纯合子分别为 33 3%、5 4 2 %和 12 5 %。结论 RDB法检测NAT2基因型准确、方便 ,适用于指导临床合理用药。 相似文献
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基于蛋白质结构的药物分子设计 总被引:2,自引:0,他引:2
随着蛋白质晶体学和分子模型技术的发展,基于蛋白质结构的药物分子设计已成为药物设计的主要潮流之一。这类方法从与疾病有关的蛋白质的三维结构出发,利用计算机程序,先对药物结合位点进行描述然后通过各种方法产生潜在的配体分子,最后对配体分子进行打争,给出设计结果。 相似文献
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黄力 《国外医学(药学分册)》2002,29(6):341-344
药用蛋白质从生物可降解聚合物中释放的过程始终影响蛋白质的三维结构、化学性质和完整性。在使用最常用的水/油/水(W/O/W)双重乳化技术包封蛋白质时,要想保持其稳定性就需要提高蛋白质的热力学稳定性或者通过使用各种添加剂使蛋白质避免接触失稳剂(如W/0界面)。然而,当人们使用W/O/W技术时,蛋白质的稳定性仍存在问题。因此,探索其他解决这种问题的方法就受到了关注。这些替代方法,如固/油/油(S/O/O)和固/油/水(S/O/W)技术是基于将干燥的蛋白质粉末悬浮于无水有机溶剂中的一种技术。很明显,蛋白质结构在有机相中的稳定性是由于蛋白质在此状态下是“僵硬的”,因而蛋白质结构的干燥是禁阻的。本文主要介绍在应用各种不同的包封技术时如何保持蛋白质结构和性能的稳定性。 相似文献
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《中国药物化学杂志》2017,(4):315-324
Sirtuin2(SIRT2)是一种烟酰胺腺嘌呤二核苷酸(NAD~+)依赖性的组蛋白去乙酰化酶,是沉默信息调节因子2(Sir2)相关蛋白家族的重要成员。SIRT2通过催化不同的底物去乙酰化参与了许多生物学过程,比如基因沉默、细胞周期调控、细胞代谢、细胞凋亡等。研究表明,在许多神经退行性疾病模型中,抑制SIRT2能够对神经元细胞产生明显的保护作用。因此,SIRT2有望成为治疗神经退行性疾病的重要靶点。本文主要对SIRT2的结构、功能、SIRT2与神经退行性疾病的关系、SIRT2抑制剂及其构效关系做一综述,旨在为以SIRT2为靶点的神经退行性疾病药物研发提供参考。 相似文献
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目的研究N-乙酰化酶2(NAT2)基因型对中国肺结核患者异烟肼(INH)代谢的影响。方法采用反向点交法(RDB)检测中国肺结核患者NAT2基因型;柱前衍生化高效液相色谱法检测服药2 h后血浆中INH及其代谢物乙酰化INH(Ac- INH)浓度。结果46例患者基因型包括野生型wt/wt(n=17)、杂合子m/wt(n=22)、突变型纯合子m/m(n=7)。血浆INH及AcINH浓度分别为(12.74±10.51)和(12.49±9.61)μmol·L~(-1)。3个基因组INH、AcINH浓度比为1:1.37:1.77和1:0.77:0.75;3组AcINH与INH比值(R_(A/I))为1:0.52:0.40(P<0.01)。结论不同NAT2基因型的结核患者对INH代谢能力差异显著,存在基因剂量现象。NAT2基因分型对结核患者合理用药具有重要意义。 相似文献
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多年来 ,对细胞色素酶P4 5 0的研究主要集中在其代谢外源性药物和毒物的方面。然而它在内源性物质 ,如类固醇、胆固醇、激素、脂肪酸和维生素等的代谢转化中也起了很重要的作用。CYP2J就是这样一种主要代谢内源性物质花生四烯酸的细胞色素酶P4 5 0超家族。人体内的CYP2J2在不久前被发现 ,它与疾病的相关性正引起研究者广泛的兴趣。本文就人类CYP2J2的组织分布、生理作用、编码基因及其基因突变等方面的研究进展作一简要综述。 相似文献
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以细菌视紫红质(bacteriorhodopsin)的三维晶体结构为模板,预测了多巴胺D2受体跨膜区的7个α螺旋肽段的三维结构。根据定点突变实验数据以及预测的受体三维结构,确定了激动剂配体结合腔由Asp86,Ser141,Ser144等12个氨基酸残基组成。为了校正和检验所得的模型,分别以一组刚性、一组柔性的激动剂与受体对接(DOCK),分析-logIC50和结合能Eb相关性,较好的结果说明该模型是可靠的。 相似文献
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《Toxicology mechanisms and methods》2013,23(8):440-444
Human arylamine N-acetyltransferase 1 (NAT1) and its homologue in rodents (Nat2) are polymorphic xenobiotic metabolizing enzymes and also seem to play a role in endogenous metabolism. NAT1 and Nat2 polymorphism was associated to cancers under xenobiotic procarcinogens metabolism as well as under endogenous substrate metabolism. This study investigated the p-aminobenzoic acid (PABA) -Nat2 catalytic activity and its polymorphism in liver homogenates of adult sand rats Psammomys obesus Cretzschmar, 1828. These Saharian sand rats develop high incidence of spontaneous cancers under standard laboratory diet. The average value of PABA-Nat2 specific activity tested in nine sand rats was significant (2.96?±?2.16 nmoles/min/mg). The N-acetylation exhibited a bimodal distribution. There was a significant difference (p?<?0.01) between PABA-Nat2 activity in the fast acetylators group (4.10?±?1.67 nmol/min/mg) and slow acetylators group (0.7?±?0.27 nmol/min/mg). The percentage of the fast acetylator group was 66.66%. These results support the presence of Nat2 polymorphism in the liver of the strain sand rats Psammomys obesus. This strain is useful for investigating the role of Nat2 polymorphisms in susceptibility to cancers related to arylamine carcinogen exposures as well as to endogenous substrate metabolism. 相似文献
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Kawamura A Westwood I Wakefield L Long H Zhang N Walters K Redfield C Sim E 《Biochemical pharmacology》2008,75(7):1550-1560
There is increasing evidence that human arylamine N-acetyltransferase type 1 (NAT1, EC 2.3.1.5), although first identified as a homologue of a drug-metabolising enzyme, appears to be a marker in human oestrogen receptor positive breast cancer. Mouse Nat2 is the mouse equivalent of human NAT1. The development of mouse models of breast cancer is important, and it is essential to explore the biological role of mouse Nat2. We have therefore produced mouse Nat2 as a recombinant protein and have investigated its substrate specificity profile in comparison with human NAT1. In addition, we have tested the effects of inhibitors on mouse Nat2, including compounds which are endogenous and exogenous steroids. We show that tamoxifen, genistein and diethylstilbestrol inhibit mouse Nat2. The steroid analogue, bisphenol A, also inhibits mouse Nat2 enzymic activity and is shown by NMR spectroscopy, through shifts in proton peaks, to bind close to the active site. A three-dimensional structure for human NAT1 has recently been released, and we have used this crystal structure to generate a model of the mouse Nat2 structure. We propose that a conformational change in the structure is required in order for ligands to bind to the active site of the protein. 相似文献
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The genotyping of N-acetyltransferase 2 (NAT2) by PCR/RFLP methods yields in a considerable percentage ambiguous results. To resolve this methodical problem a statistical
approach was applied. PHASE v2.1.1, a statistical program for haplotype reconstruction was used to estimate haplotype pairs
from NAT2 genotyping data, obtained by the analysis of seven single nucleotide polymorphisms relevant for Caucasians. In 1,011 out
of 2,921 (35%) subjects the haplotype pairs were clearcut by the PCR/RFLP data only. For the majority of the data the applied
method resulted in a multiplicity (2–4) of possible haplotype pairs. Haplotype reconstruction using PHASE v2.1.1 cleared this
ambiguity in all cases but one, where an alternative haplotype pair was considered with a probability of 0.029. The estimation
of the NAT2 haplotype is important because the assignment of the NAT2 alleles *12A, *12B, *12C or *13 to the rapid or slow NAT2 genotype has been discussed controversially. A clear assignment is indispensable in surveys of human bladder cancer caused
by aromatic amine exposures. In conclusion, PHASE v2.1.1 software allowed an unambiguous haplotype reconstruction in 2,920
of 2,921 cases (>99.9%). 相似文献
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Straka RJ Burkhardt RT Lang NP Vang T Hadsall KZ Tsai MY 《Biopharmaceutics & drug disposition》2006,27(6):299-304
Southeast Asians known as the Hmong have a high prevalence of tuberculosis and select cancers. The slow acetylation (SA) phenotype for N-acetyltransferase 2 (NAT2) has been associated with toxicity from the anti-tuberculosis drug, isoniazid and in increased risk of select cancers. Previous research indicates a 74.5% prevalence of SA in Hmong which differs from other Asian populations including the Japanese and Thai (range: 7%-45%). Given this contrast, the purpose of this study was to confirm or refute this unexpected predominance of the SA phenotype in Hmong. Unrelated, Minnesota Hmong between 18 and 65 years of age consented and participated by ingesting caffeine as the probe for NAT2. A urinary caffeine metabolic ratio AFMU/1X (<0.6) was used to classify subjects as slow acetylators. Among 51 analysable samples provided by 61 enrollees (27 male, 33 female, 1 sex unknown, age 30+/-11 years [mean+/-SD]) there were 47 (92.2%) slow and 4 (7.8%) rapid acetylators. The prevalence of the SA phenotype (92.2%) from this study exceeds the 74.5% (p<0.02 by chi-square test) previously noted in Minnesota Hmong (n=98). The predominance of the SA phenotype within Minnesota Hmong is confirmed. Further studies evaluating this unexpected prevalence, its genetic basis and potential clinical relevance to drug toxicity and disease are warranted. 相似文献
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Inactivation of human arylamine N-acetyltransferase 1 by the hydroxylamine of p-aminobenzoic acid 总被引:1,自引:0,他引:1
Human N-acetyltransferase 1 (NAT1) is a widely distributed enzyme that catalyses the acetylation of arylamine and hydrazine drugs as well as several known carcinogens, and so its levels in the body may have toxicological importance with regard to drug toxicity and cancer risk. Recently, we showed that p-aminobenzoic acid (PABA) was able to down-regulate human NAT1 in cultured cells, but the exact mechanism by which PABA acts remains unclear. In the present study, we investigated the possibility that PABA-induced down-regulation involves its metabolism to N-OH-PABA, since N-OH-AAF functions as an irreversible inhibitor of hamster and rat NAT1. We show here that N-OH-PABA irreversibly inactivates human NAT1 both in cultured cells and cell cytosols in a time- and concentration-dependent manner. Maximal inactivation in cultured cells occurred within 4 hr of treatment, with a concentration of 30 μM reducing activity by 60 ± 7%. Dialysis studies showed that inactivation was irreversible, and cofactor (acetyl coenzyme A) but not substrate (PABA) completely protected against inactivation, indicating involvement of the cofactor-binding site. In agreement with these data, kinetic studies revealed a 4-fold increase in cofactor Km, but no change in substrate Km for N-OH-PABA-treated cytosols compared to control. We conclude that N-OH-PABA decreases NAT1 activity by a direct interaction with the enzyme and appears to be a result of covalent modification at the cofactor-binding site. This is in contrast to our findings for PABA, which appears to reduce NAT1 activity by down-regulating the enzyme, leading to a decrease in NAT1 protein content. 相似文献
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Reema Deol 《Xenobiotica; the fate of foreign compounds in biological systems》2017,47(3):202-207
1.?The mercapturic acid (MA) pathway is a metabolic route for the processing of glutathione conjugates to MA (N-acetylcysteine conjugates). An N-acetyltransferase enzyme, NAT8, catalyzes the transfer of an acetyl group from acetyl-CoA to the cysteine amino group, producing a MA, which is excreted in the urine. We expressed human NAT8 in HEK293T cells and developed an HPLC-MS method for the quantitation of the S-aryl-substituted cysteine conjugates and their MA.2.?We measured the activity of the enzyme for acetylation of benzyl-, 4-nitrobenzyl-, and 1-menaphthylcysteine substrates.3.?NAT8 catalyzed the acetylation of all three cysteine conjugates with similar Michaelis–Menten kinetics. 相似文献
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甲状腺乳头状癌的治疗预后虽然较好,但是同样具有恶性肿瘤侵袭、转移的生物学特性。MMPs在肿瘤的侵袭、转移过程中发挥了重要的作用,尤其是其家族成员中的MMP-2(基质金属蛋白酶-2或明胶酶)的研究,提示在甲状腺乳头状癌组织中呈高阳性,高水平表达与良性病变有明显的统计学差异。本文结合近年来对MMP-2在甲状腺疾病中研究的国内外报道文献,综述MMP-2在甲状腺乳头状癌的研究状况,展望未来的研究方向。 相似文献
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NAT2基因多态性与柳氮磺吡啶治疗溃疡性结肠炎安全性的关系 总被引:2,自引:0,他引:2
目的:探讨N-乙酰基转移酶2(NAT2)基因型多态性与柳氮磺吡啶(SASP)治疗溃疡性结肠炎(UC)安全性的关系。方法:在38例UC患者及36例健康对照者中,采用聚合酶链式反应-限制性片段长度多态性(PCR—RFLP)法检测NAT2的基因多态型,并对UC患者进行NAT2基因型与SASP药物不良反应(ADR)的相关性分析。结果:慢型乙酰化基因型的UC患者服用SASP后ADR的发生率高于快型乙酰化基因型患者,差异有统计学意义(P〈0.01)。结论:UC患者使用SASP出现的ADR与NAT2慢型乙酰化基因型相关。 相似文献