JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
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2022
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IF 3.318
Xuan Guo,Lei Zhang,Zihan Lei,Zhe Hou,Hui Li,Xiaodong Li,Jing Dong,Ling Song,Dingding Chen,Dongyang Liu
Objective The aim of this study is to develop a LC-MS/MS method for the quantitation of seven cytochrome P450 (CYP450) enzymes. Methods A high-performance liquid chromatography-tandem mass spectrometry method was developed using multiple reaction monitoring mode with positive electrospray ionization. The method was validated with selectivity, linearity, stability, accuracy and precious. In addition, the abundance of seven CYP450 enzymes in human liver microsomes and CYP3A4 in placenta were determined using the current method. Results The linear range for CYP1A2, CYP2B6 and CYP2C8 was 0.036–3.6 nM and for CYP2C9, CYP2C19, CYP2D6 and CYP3A4 was 0.090–9.0 nM. No interference was found between the blank matrix and each specific peptides. The accuracy and precious results were in accord with the requirement of analytical methods for biological samples in Chinese Pharmacopoeia. In addition, the peptides were stable under current stability conditions. The content of CYP3A4 in placenta and the seven CYP450 enzymes in human liver microsomes were accurately quantified. Conclusion The developed method is sensitive and specific and can be applied to the quantification of enzymes abundance in different human derived samples like placenta and liver microsomes.
研究领域:
药理学
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分析化学
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生物化学
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DOI
PubMed
高影响因子
PESTICIDE BIOCHEMISTRY AND PHYSIOLOGY
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2022
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IF 4.966
Bin Liu,Danyang Fu,Hang Ning,Ming Tang,Hui Chen
Bark beetles rely on detoxifying enzymes to resist the defensive terpenoids of the host tree. Insect cytochrome P450 (CYPs) plays a key role in the detoxification of pesticides and plant allelochemicals . CYP6 family is unique to Insecta, and its biochemical function is basically related to the metabolism of exogenous substances. In this study, we sequenced and characterized the full-length cDNAs of two CYP6 genes from Chinese white pine beetle, Dendroctonus armandi . Spatiotemporal expression profiling revealed that the expression of CYP6CR2 and CYP6DE5 was higher in larval and adult stages of D. armandi than that in other developmental stages, and that two genes predominantly expressed in brain , midgut, fat body, Malpighian tubules or hemolymph. The expression of CYP6CR2 and CYP6DE5 was significantly induced after feeding on the phloem of Pinus armandii and exposure to six stimuli [(±)- α -pinene, (−)- α -pinene, (−)- β -pinene, (+)-3-carene, (±)-limonene and turpentine]. Importantly, silencing CYP6CR2 and CYP6DE5 separately could increase the sensitivity, led to a significant reduction of the activity of P450, resulting a significant increase in adult mortality after treatment with terpenoids. The comprehensive results of this study showed that in the process of host selection and colonization, the functions of CYPs were mainly to hydrolyze the chemical defense of the host and degrade odor molecules. These findings may help to develop new treatments to control this important pest.
研究领域:
昆虫学
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分子生物学
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生物化学
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DOI
PubMed
高影响因子
PEST MANAGEMENT SCIENCE
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2025
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IF 3.8
Juncheng Wu,Zhengyuan Tao,Jingyi Cao,Wei Hu,Minghao Jiang,Yahui Li,Haiqun Cao,Min Liao,Ning Zhao
BACKGROUND Rice flatsedge ( Cyperus iria L.) is one of the most troublesome weeds infesting rice fields across China. Bensulfuron-methyl, an acetolactate synthase (ALS)-inhibiting herbicide, has been widely used for the control of Cyperaceae weeds in rice production. However, long-term and extensive use of this herbicide has resulted in the evolution of resistant C. iria populations. In this study, a suspected bensulfuron-methyl-resistant (R) population collected from a rice field that survived field-recommended applications was investigated to elucidate its resistance level and underlying mechanism. RESULTS Compared with a susceptible (S) population, the R population exhibited a high level of resistance to bensulfuron-methyl [resistance index (RI) = 12.88] and cross-resistance to metazosulfuron (RI = 11.66), bispyribac-sodium (RI = 9.10) and penoxsulam (RI = 6.35). No mutations were detected in the ALS gene, and ALS expression levels did not differ significantly between the R and S plants. Pretreatment with the cytochrome P450 inhibitor malathion and the glutathione S -transferase inhibitor 4-chloro-7-nitrobenzoxadiazole effectively reversed bensulfuron-methyl resistance in R plants. Liquid chromatography tandem mass spectrometry analysis showed that the R plants metabolized bensulfuron-methyl significantly faster than the S plants. RNA sequenccing analysis revealed remarkable upregulation of CYP97A3 and GSTF1 in the R population, while molecular docking indicated strong binding affinities between both enzymes and bensulfuron-methyl at their active sites. CONCLUSION These results reveal that enhanced expression of CYP97A3 and GSTF1 may contribute to bensulfuron-methyl resistance in C. iria , highlighting the role of metabolic detoxification in the evolution of non-target-site resistance in this species. © 2025 Society of Chemical Industry.
研究领域:
杂草科学
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植物生理学
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分子生物学
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除草剂抗性
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农业植物学
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DOI
PubMed
高影响因子
BIOCHEMICAL PHARMACOLOGY
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2026
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IF 6.5
Junze Huang,Luyao Deng,Bingyi Yao,Luping Zhao,Xin Xu,Lei Liu,Chenmeizi Liang,Yan Shen,Yuanjin Zhang,Xin Wang
Progesterone (PR) is an essential steroid hormone regulating female reproductive function, whereas fluoxetine (FLU), a selective serotonin reuptake inhibitor (SSRI), serves as a first-line antidepressant for antenatal use. The two agents may undergo cytochrome P450 (CYP)-mediated interactions, threatening maternal and fetal safety. This study integrated cocktail incubation, in vitro / in vivo assays, and mechanistic static models (MSM) to characterize PR-modulated CYP activity and FLU-associated drug-drug interactions (DDI). In human/rat liver and human placental microsomes, PR exerted reversible competitive inhibition on human hepatic CYP2C9 (IC 50 = 26.16 ± 3.89 μM, K i = 34.64 μM) and rat CYP2C11 (IC 50 = 20.81 ± 3.49 μM, K i = 38.30 μM), with no inhibitory effect on placental CYPs. It suppressed FLU metabolism in rat liver via mixed inhibition (IC 50 = 16.56 ± 1.50 μM). In vivo , intramuscular progesterone (IMP) elevated rat FLU AUC 0-t by 57.4% and C max by 123.7%, while lowering CL/F by 42.1%. MSM predicted evident perinatal DDI risks for high-dose IMP (AUCR up to 1.53), driven by altered hepatic first-pass metabolism. Though the general population faces low risks, close therapeutic monitoring is mandatory for pregnant women. This work underscores the delivery route, physiological status, and patient subgroups in DDI evaluation, supporting the safe combined use of PR and CYP2C9 substrates.
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高影响因子
JOURNAL OF BIOLOGICAL CHEMISTRY
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2026
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IF 4.1
Yunjie Liu,Yue Zhao,Weihan Sun,Nian Li,Yunjun Pan,Li Ma,Shengying Li
Steroids with anti-inflammatory, anti-allergic, endocrine-regulating, and other pharmaceutical activities represent the second most widely used class of drugs worldwide, following antibiotics. Their industrial production primarily relies on Mycobacteria -mediated biotransformation of sterols into key intermediates, followed by chemical or enzymatic modifications. While the sterol metabolic pathways in Mycobacteria have been intensively studied, the identification and functional characterization of key enzymes, particularly cytochrome P450 enzymes (CYPs or P450s) and their cognate redox partners, remain incomplete. Here, we heterologously expressed 24 P450s, 10 ferredoxin reductases (FdRs), and 12 ferredoxins (Fdxs) from Mycobacterium neoaurum ZC-1 in Escherichia coli . In vitro biochemical experiments identified five P450 enzymes (CYP125A76, CYP125A77, CYP125A78, CYP142A12, and CYP124A1) capable of catalyzing sterol side-chain terminal oxidation. Screening 120 redox partner combinations revealed FdR4662/Fdx4443 as the optimal cognate redox partners for all five P450 enzymes. With this redox partner pair, CYP142A12 achieved a conversion ratio of 89 % for 4-cholesten-3-one with NADH as the preferred cofactor. Structural analyses indicate that the electron-transfer efficiency is primarily governed by electrostatic complementarity around the Fe–S cluster, the redox-center distance between the Fe–S cluster and heme-iron, and the FAD-to-cluster distance within the FdR-Fdx complex. These findings highlight the critical role of redox partner selection in enhancing P450 catalytic efficiency and provide a solid foundation for engineering high-efficiency industrial strains to improve steroid biomanufacturing and reduce production costs.
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DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
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2026
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IF 8.7
Chao Xiang,Jia-Jia Mou,Yue Jiang,Yu-Ke Cen,Ya-Ping Xue,Yu-Guo Zheng
Cytochrome P450 enzymes hold immense potential as biocatalysts for oxidative reactions but often suffer from inefficient electron transfer due to their reliance on transient interactions with redox partners (RPs). To overcome this limitation, we constructed self-sufficient fusion enzymes by fusing the avermectin-oxidizing P450-Ema1-M212A to selected RPs. Linker sequences were primarily optimized experimentally, while computational modeling, including AlphaFold3-predicted multi-protein assemblies and molecular dynamics simulations, was used as an auxiliary tool to assess feasible linker lengths and interpret the structural basis by which optimized linker design enhances electron-transfer efficiency. Based on literature precedent and preliminary structural considerations, a series of fusion constructs with varying linker sequences and lengths was constructed. Molecular dynamics simulations suggested a structural rationale for why a linker length of 9 amino acids exhibited optimal enzymatic activity, as it stabilized a functional conformation, minimized structural fluctuations, and maintained a favorable distance between the heme and Fe₂S₂ clusters for efficient electron transfer. Experimental characterization confirmed that the F4 variant exhibited substantially increased activity over the non-fusion version. Kinetic analysis demonstrated a significantly enhanced k cat / K m ratio, indicating accelerated catalysis without compromising substrate binding. For the synthesis of 4″-oxo-avermectin, a key intermediate of emamectin, the F4 variant achieved 90% conversion within 8 h, far exceeding the performance of the non-fused system. This study establishes an experimentally driven, semi-rational framework for constructing self-sufficient P450 fusion enzymes, highlighting the synergy between computational prediction and protein engineering for developing powerful biocatalysts with high industrial application potential.
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PubMed
高影响因子
Nature Communications
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2026
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IF 18.1
Jiang Yuanyuan,Zhang Junjie,Li Zhong,Wang Lingyan,Guo Yuyu,Li Yuxuan,Feng Xisong,Gao Wen,Li Yong,Li Zijia,Du Jieke,Zhang Guoqiang,Gong Piqian,Fang Wenhan,Gao Xiang,Bai Mingyi,Hollmann Frank,Zhang Mei,Huang He,Wang Binju,Mo Sudong,Li Huarong,Peng Wei,Li Shengying
Herbicides in combination with genetically modified herbicide-resistant crops have revolutionized modern weed management, increased crop yields, and facilitated farming practices. However, rapid evolution of herbicide-resistant weeds necessitates new resistance traits to sustain control efficacy. Here, we introduce a terminal carboxyl anchoring mechanism-inspired approach for precise discovery of P450 herbicide resistance genes, by which a number of bacterial P450 peroxygenases are predicted and confirmed to degrade auxin herbicides. Upon enzyme engineering, the optimal mutant P450 BSβ -F46A can efficiently degrade diverse auxin herbicides and other carboxyl-containing herbicides. Mechanistic studies reveal that Compound I-mediated hydroxylation initiates the C‒O bond cleavage, followed by aromatic ring hydroxylation, thus forming a unique two-step degradation pathway. Transgenic rice expressing P450 BSβ -F46A-CPR confers significant resistance to a recently commercialized auxin herbicide fluchloraminopyr. This work demonstrates the potential of the mechanism-driven strategy in directed discovery of broad-spectrum resistance genes for herbicide-resistant crop engineering.
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DOI
高影响因子
Nature Communications
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2025
·Vol. 16
·期 1
·1-18
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IF 15.7
Li Wanqian,Whiteway Malcolm,Hang Sijin,Yu Jinhua,Lu Hui,Jiang Yuanying
Azole effectiveness against candidiasis can be compromised by Candida albicans resistance and tolerance, and unfortunately, few clinically useful compounds can enhance azole antifungal activities. We find that the amino acids V234, F235 and L238 of Erg11 are critical for its interaction with Ncp1, and the Ncp1-Erg11 association is important in azole response. Ellipticine and its analog phiKan 083 block this Erg11-Ncp1 interaction by targeting Ncp1, and boost antifungal effects of fluconazole in vitro and in vivo. A series of steps influencing this process—an initial elevation in reactive oxygen species, leading to protein oxidation and misfolding in the endoplasmic reticulum (ER) that causes ER stress. This stress leads to Ca 2+ release from the ER, mitochondrial Ca 2+ accumulation and dysfunction, increased ROS production, and apoptosis of C. albicans cells. Overall, disrupting the Erg11-Ncp1 interaction in C. albicans can serve as a useful approach to enhancing the antifungal properties of azoles.
研究领域:
微生物学
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分子生物学
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药理学
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生物化学
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医学真菌学
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产品关联
DOI
PubMed
高影响因子
ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY
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2026
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IF 6.6
Yue Li,Zengcui Yang,Lijuan Luo,Ling Fang,Xinyan Li,Shanshan Chen,Yaobin Lu,Tiangang Luan
Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental contaminants which pose significant risks to ecological environments and human health. The microalga Raphidocelis subcapitata demonstrates a remarkable capacity for PAH degradation. Given that PAHs with different molecular weights exhibit distinct physicochemical properties which may affect the activity of PAHs-degrading enzymes within microalgae. Transcriptomic analysis of R. subcapitata exposed to three PAHs – phenanthrene (PHE), pyrene (PYR), and benzo[ a ]pyrene (BAP) – identified a total of 14,075 differentially expressed genes across all treatments. Three genes encoding glutathione S-transferase ( gSt ) and two cytochrome P450 isoforms ( CYPA and CYPB ) were further validated by RT-qPCR; all of which are functionally associated with PAH biodegradation in R. subcapitata . Treatment with 1-aminobenzotriazole (ABT), a cytochrome P450 (CYP450) enzyme inhibitor, decreased the microalgal degradation of PHE, PYR, and BAP by 86.0%, 72.2%, and 48.3%, respectively, relative to the ABT-free algal control. Specifically, ABT primarily inhibited the formation of monohydroxylated metabolites of PHE and PYR, suggesting that monooxygenases were the primary metabolic enzymes responsible for the degradation of PHE and PYR, while the degradation of BAP likely involved a collaborative action between monooxygenases and dioxygenases. This study provided new evidence for the mechanism of microalgal PAH degradation and theoretical support for the application of R. subcapitata in remediating PAH-contaminated freshwater environments.
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高影响因子
PHYTOMEDICINE
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2017
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IF 3.526
Jian Lu,Daozhi Liu,Xiaojing Zhou,Ang Chen,Zhenran Jiang,Xiyun Ye,Mingyao Liu,Xin Wang
Background Cytochrome P450 2J2 (CYP2J2) is not only highly expressed in many kinds of human tumors, but also promotes tumor cell growth via regulating the metabolism of arachidonic acids . CYP2J2 inhibitors can significantly reduce proliferation, migration and promote apoptosis of tumor cells by inhibiting epoxyeicosatrienoic acids (EETs) biosynthesis . Therefore screening CYP2J2 inhibitors is a significant way for the development of anti-cancer drug. Purpose The aim of this study was to identify a new CYP2J2 inhibitor from fifty natural compounds obtained from plants. Study design CYP2J2 inhibitor was screened from a natural compounds library and further the inhibitory manner and mechanism were evaluated. Its cytotoxicity against HepG2 and SMMC-7721 cell lines was also estimated. Methods The inhibitory effect was evaluated in rat liver microsomes (RLMs), human liver microsomes (HLMs) and recombinant CYP2J2 (rCYP2J2), using astemizole as a probe substrate and inhibitory mechanism was illustrated through molecular docking . The cytotoxicity was detected using SRB. Results In all candidates, plumbagin showed the strongest inhibitory effect on the CYP2J2-mediated astemizole O -demethylation activity. Further study revealed that plumbagin potently inhibited CYP2J2 activity with IC 50 value at 3.82 µM, 3.37 µM and 1.17 µM in RLMs, HLMs and rCYP2J2, respectively. Enzyme kinetic studies showed that plumbagin was a mixed-type inhibitor of CYP2J2 in HLMs and rCYP2J2 with K i value of 1.88 µM and 0.92 µM, respectively. Docking data presented that plumbagin interacted with CYP2J2 mainly through GLU 222 and ALA 223. Moreover, plumbagin showed strongly cytotoxic effects on hepatoma cell lines, such as HepG2 and SMMC-7721, with lower toxicity on rat primary hepatocytes. Plumbagin had no effect on the protein expression of CYP2J2 in HepG2 and SMMC-7721, while down-regulated the mRNA level of anti-apoptosis protein Bcl-2. Conclusion This study found out a new CYP2J2 inhibitor plumbagin from fifty natural compounds. Plumbagin presented a potential of anti-cancer pharmacological activity .
研究领域:
药理学
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肿瘤学
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分子生物学
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DOI
PubMed
高影响因子
MOLECULAR CARCINOGENESIS
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2018
·Vol. 57
·期 10
·1371-1382
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IF 3.851
Jie Gao,Zhao Wang,Gao-Ju Wang,Na Gao,Jing Li,Yun-Fei Zhang,Jun Zhou,Hong-Xin Zhang,Qiang Wen,Han Jin,Hai-Ling Qiao
Hepatofibrosis is an important susceptibility factor for hepatocarcinogenesis. However, only a handful of cases of hepatofibrosis will develop into hepatocellular carcinoma (HCC). As cytochrome P450 2E1 (CYP2E1) is involved in the metabolism and activation of many known environmental toxicants and procarcinogens, this enzyme may play a role in the development of hepatocarcinogenesis subsequent to hepatofibrosis. Herein, we evaluated whether higher CYP2E1 activity is a risk factor for the development of hepatocarcinogenesis from hepatofibrosis. CYP2E1 activity in fibrotic tissues from 72 HCC patients and in normal liver tissues from 59 control subjects was determined along with the severity of hepatofibrosis in hepatocarcinogenesis patients. Similarly, using a rat diethylnitrosamine-induced hepatocarcinogenesis model, CYP2E1 activity at the hepatofibrosis and hepatocarcinogenesis stages was determined, the correlation between CYP2E1 activity at the hepatofibrosis stage and hepatocarcinogenesis was explored, and the impact of inhibition of CYP2E1 activity on hepatocarcinogenesis was studied. The results showed that while CYP2E1 activity in HCC patients with underlying hepatofibrosis was increased, the severity of hepatofibrosis did not correlate with CYP2E1 activity. In the rat hepatocarcinogenesis model, unexpectedly, CYP2E1 activity was found to decrease from hepatofibrosis to hepatocarcinogenesis. Importantly, however, hepatofibrotic rats with higher CYP2E1 activity developed a more severe form of HCC. Moreover, inhibition of CYP2E1 activity could decrease the occurrence and development of HCC in rats. In conclusion, higher CYP2E1 activity may be a risk factor for hepatocarcinogenesis from hepatofibrosis, which raises the possibility of screening patients with hepatofibrosis for CYP2E1 activity to better estimate their risk for hepatocarcinogenesis.
研究领域:
医学
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药理学
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毒理学
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DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
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2023
·Vol. 24
·期 3
·2395
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IF 5.6
Yanqing Li,Yingwu Lin,Fang Wang,Jinghan Wang,Osami Shoji,Jiakun Xu
With the increasing demand for blue dyes, it is of vital importance to develop a green and efficient biocatalyst to produce indigo. This study constructed a hydrogen peroxide-dependent catalytic system for the direct conversion of indole to indigo using P450BM3 with the assistance of dual-functional small molecules (DFSM). The arrangements of amino acids at 78, 87, and 268 positions influenced the catalytic activity. F87G/T268V mutant gave the highest catalytic activity withkcatof 1402 min−1and with a yield of 73%. F87A/T268V mutant was found to produce the indigo product with chemoselectivity as high as 80%. Moreover, F87G/T268A mutant was found to efficiently catalyze indole oxidation with higher activity (kcat/Km= 1388 mM−1min−1) than other enzymes, such as the NADPH-dependent P450BM3 (2.4-fold), the Ngb (32-fold) and the Mb (117-fold). Computer simulation results indicate that the arrangements of amino acid residues in the active site can significantly affect the catalytic activity of the protein. The DFSM-facilitated P450BM3 peroxygenase system provides an alternative, simple approach for a key step in the bioproduction of indigo.
研究领域:
生物技术
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酶工程
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绿色化学
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）