Please wait a minute...
Journal of Integrative Agriculture  2020, Vol. 19 Issue (6): 1512-1521    DOI: 10.1016/S2095-3119(19)62827-3
Special Focus: Physiology and interaction of insects with environmental factors Advanced Online Publication | Current Issue | Archive | Adv Search |
Characteristics and roles of cytochrome b5 in cytochrome P450-mediated oxidative reactions in Locusta migratoria
LIU Jiao1, 2, ZHANG Xue-yao1, WU Hai-hua1, MA Wen3, ZHU Wen-ya4, Kun-Yan ZHU5, MA En-bo1, ZHANG Jian-zhen1 
1 Institute of Applied Biology, Shanxi University, Taiyuan 030006, P.R.China
2 College of Life Science, Shanxi University, Taiyuan 030006, P.R.China
3 Modern Research Center for Traditional Chinese Medicine, Shanxi University, Taiyuan 030006, P.R.China
4 Institute of Plant Protection, Shanxi Academy of Agricultural Sciences, Taiyuan 030031, P.R.China
5 Department of Entomology, Kansas State University, Manhattan, KS 66506, USA
Download:  PDF in ScienceDirect  
Export:  BibTeX | EndNote (RIS)      
Cytochrome b5 (Cyt-b5) is a small heme protein and known to be involved in a wide range of biochemical transformations, including cytochrome P450 monooxygenase (CYP)-mediated metabolism of endogenous and exogenous compounds.  Studies on Cyt-b5 are more concentrated in mammals, but are relatively rare in insects.  The characteristics and function of Cyt-b5 from Locusta migratoria have not been described yet.  We sequenced the full-length cDNA sequence of Cyt-b5 from L. migratoria (LmCyt-b5) by reverse transcription-PCR (RT-PCR) based on locust transcriptome database.  The phylogenetic analysis showed that LmCyt-b5 was closely related to the Cyt-b5 from Blattodea.  LmCyt-b5 was highly expressed in ovary, Malpighian tubules, midgut, gastric caeca, and fat bodies.  Silencing of LmCyt-b5 had no effect on the susceptibility of L. migratoria to four different insecticides.  Suppression of LmCyt-b5 or silencing of both LmCyt-b5 and LmCPR did not significantly change the total CYP activity toward the substrate 7-ethoxycoumarin (7-EC).  However, coexpression of LmCYP6FD1 with LmCPR and LmCyt-b5 together in Sf9 cells by using Bac-to-Bac baculovirus expression system significantly increased the catalytic activity of LmCYP6FD1 toward 7-EC as compared with the coexpression of LmCYP6FD1 with cytochrome P450 reductase (LmCPR) or LmCyt-b5 separately.  These results suggest that LmCyt-b5 plays an important role in the catalytic reaction of LmCYP6FD1 toward 7-EC in our in vitro experiments.  Further study is needed to clarify the role of LmCyt-b5 in CYP-mediated catalytic reactions in L. migratoria.
Keywords:  cytochrome b5        cytochrome P450        cytochrome P450 reductase        Locusta migratoria        RNA interference  
Received: 14 May 2019   Accepted: 26 April 2020
Fund: This research was supported by the National Natural Science Foundation of China (31320103921 and 31872010) and the Graduate Outstanding Innovation Project of Shanxi Province, China (2017 BY011).
Corresponding Authors:  Correspondence MA En-bo, Tel: +86-351-7018871, E-mail:; ZHANG Jian-zhen, Tel: +86-351-7018871, E-mail:   
About author:  LIU Jiao, Mobile: +86-15536053652, E-mail:;

Cite this article: 

LIU Jiao, ZHANG Xue-yao, WU Hai-hua, MA Wen, ZHU Wen-ya, Kun-Yan ZHU, MA En-bo, ZHANG Jian-zhen . 2020. Characteristics and roles of cytochrome b5 in cytochrome P450-mediated oxidative reactions in Locusta migratoria. Journal of Integrative Agriculture, 19(6): 1512-1521.

Bart A G, Scott E E. 2017. Structural and functional effects of cytochrome b5 interactions with human cytochrome P450 enzymes. Journal of Biological Chemistry, 292, 20818–20833.
Chen X, Zhang Y. 2015. Identification and characterization of NADPH-dependent cytochrome P450 reductase gene and cytochrome b5 gene from Plutella xylostella: Possible involvement in resistance to beta-cypermethrin. Gene, 558, 208–214.
Feyereisen R. 2011. Arthropod CYPomes illustrate the tempo and mode in P450 evolution. Biochimica et Biophysica Acta (Proteins and Proteomics), 1814, 19–28.
Feyereisen R. 2012. Insect CYP genes and P450 enzymes. In: Gilbert L I, ed., Insect Molecular Biology and Biochemistry. Elsevier Academic Press, Oxford, UK. pp. 236–316.
Fukushima H, Grinstead G F, Gaylor J L. 1981. Total enzymic synthesis of cholesterol from lanosterol cytochrome b5-dependence of 4-methyl sterol oxidase. Journal of Biological Chemistry, 256, 4822–4826.
Guo W, Wang X, Ma Z, Xue L, Han J, Yu D, Kang L. 2011. CSP and Takeout genes modulate the switch between attraction and repulsion during behavioral phase change in the migratory locust. PLoS genetics, 7, e1001291.
Guo Y, Wu H, Zhang X, Ma E, Guo Y, Zhu K Y, Zhang J. 2016. RNA interference of cytochrome P450 CYP6F subfamily genes affects susceptibility to different insecticides in Locusta migratoria. Pest Management Science, 72, 2154–2165.
Guo Y, Zhang X, Wu H, Yu R, Zhang J, Zhu K Y, Guo Y, Ma E. 2015. Identification and functional analysis of a cytochrome P450 gene CYP9AQ2 involved in deltamethrin detoxification from Locusta migratoria. Pesticide Biochemistry and Physiology, 122, 1–7.
Guo Y, Zhang J, Yu R, Zhu K Y, Guo Y, Ma E. 2012. Identification of two new cytochrome P450 genes and RNA interference to evaluate their roles in detoxification of commonly used insecticides in Locusta migratoria. Chemosphere, 87, 709–717.
Hatakeyama M, Kitaoka T, Ichinose H. 2016. Heterologous expression of fungal cytochromes P450 (CYP5136A1 and CYP5136A3) from the white-rot basidiomycete Phanerochaete chrysosporium: Functionalization with cytochrome b5 in Escherichia coli. Enzyme and Microbial Technology, 89, 7–14.
Henderson C J, McLaughlin L A, Scheer N, Stanley L A, Wolf C R. 2015. Cytochrome b5 is a major determinant of human cytochrome P450 CYP2D6 and CYP3A4 activity in vivo. Molecular Pharmacology, 87, 733–739.
Kandel S E, Lampe J N. 2014. Role of protein-protein interactions in cytochrome P450-mediated drug metabolism and toxicity, Chemical Research in Toxicology, 27, 1474–1486.
Karunker I, Morou E, Nikou D, Nauen R, Sertchook R, Stevenson B J, Paine M J I, Morin S, Vontas J. 2009. Structural model and functional characterization of the Bemisia tabaci CYP6CM1vQ, a cytochrome P450 associated with high levels of imidacloprid resistance. Insect Biochemistry and Molecular Biology, 39, 697–706.
Kawano T, Kozutsumi, Y, Kawasaki T, Suzuki A. 1994. Biosynthesis of N-glycolylneuraminic acid-containing glycoconjugates. Purification and characterization of the key enzyme of the cytidine monophospho-N-acetylneuraminic acid hydroxylation system. Journal of Biological Chemistry, 269, 9024–9029.
Keyes S R, Cinti D L. 1980. Biochemical properties of cytochrome b5-dependent microsomal fatty acid elongation and identification of products, Journal of Biological Chemistry, 255, 11357–11364.
Laursen T, Jensen K, Møller B L. 2011. Conformational changes of the NADPH-dependent cytochrome P450 reductase in the course of electron transfer to cytochromes P450. Biochimica et Biophysica Acta (Proteins and Proteomics), 1814, 132–138.
Lee K B, La Mar G N, Kehres L A, Fujinari E M, Smith K M, Pochapsky T C, Sligar S G. 1990. Proton NMR study of the influence of hydrophobic contacts on protein prosthetic group recognition in bovine and rat ferri cytochrome b5. Biochemistry, 29, 9623–9631.
Li X, Schuler M A, Berenbaum M R. 2007. Molecular mechanisms of metabolic resistance to synthetic and natural xenobiotics. Annual Review of Entomology, 52, 231–253.
Liu J, Wu H, Zhang X, Ma W, Zhu W, Silver K, Ma E, Zhang J, Zhu K Y. 2019. Metabolism of selected model substrates and insecticides by recombinant CYP6FD1 encoded by its gene predominately expressed in the brain of Locusta migratoria. Pesticide Biochemistry and Physiology, 159, 154–162.
Liu J, Zhang X, Wu H, Gao Y, Silver K, Ma E, Zhang J, Zhu K Y. 2018. Comparisons of microsomal cytochrome P450 content and enzymatic activity towards selected model substrates and insecticides in different tissues from the migratory locust (Locusta migratoria). Chemosphere, 208, 366–373.
Livak K J, Schmittgen T D. 2001. Analysis of relative gene expression data using real time quantitative PCR and the 2–??CT method. Methods, 25, 402–408.
Masters B S S, Okita R T. 1980. The history, properties, and function of NADPH cytochrome P-450 reductase. Pharmacology & Therapeutics Pharmacol, 9, 227–244.
Mathews F S. 1985. The structure, function and evolution of cytochromes. Progress in Biophysics and Molecular Biology, 45, 1–56.
Murataliev M B, Guzov V M, Walker F A, Feyereisen R. 2008. P450 reductase and cytochrome b5 interactions with cytochrome P450: Effects on house ?y CYP6A1 catalysis. Insect Biochemistry and Molecular Biology, 38, 1008–1015.
Paltauf F, Prough R A, Masters B S S, Johnston J M. 1974. Evidence for the participation of cytochrome b5 in plasmalogen biosynthesis. Journal of Biological Chemistry, 249, 2661–2662.
Pearl N M, Wilcoxen J, Im S, Kunz R, Darty J, Britt R D, Ragsdale S W, Waskell L. 2016. Protonation of the hydroperoxo intermediate of cytochrome P450 2B4 is slower in the presence of cytochrome P450 reductase than in the presence of cytochrome b5. Biochemistry, 55, 6558–6567.
Ranasinghe C, Hobbs A A. 1999. Isolation and characterisation of a cytochrome b5 cDNA clone from Helicoverpa armigera (Hübner): possible involvement of cytochrome b5 in cytochrome P450 CYP6B7 activity towards pyrethroids. Insect Biochemistry and Molecular Biology, 29,145.
Schenkman J B, Jansson I. 2003. The many roles of cytochrome b5. Pharmacology & Therapeutics, 97, 139–152.
Sezutsu H, Le Goff G, Feyereisen R. 2013. Origins of P450 diversity. Philosophical Transactions of the Royal Society (B: Biological Sciences), 368, 20120428.
Stevenson B J, Bibby J, Pignatelli P, Muangnoicharoen S, O’Neill P M, Lian L, Müller P, Nikou D, Steven A, Hemingway J, Sutcliffe M J, Paine M J. 2011. Cytochrome P450 6M2 from the malaria vector Anopheles gambiae metabolizes pyrethroids: Sequential metabolism of deltamethrin revealed. Insect Biochemistry and Molecular Biology, 41, 492–502.
Stiborová M, Indra R, Frei E, Kope?ková K, Schmeiser H H, Eckschlager T, Adam V, Heger Z, Arlt V M, Martínek V. 2017. Cytochrome b5 plays a dual role in the reaction cycle of cytochrome P450 3A4 during oxidation of the anticancer drug ellipticine. Monatshefte für Chemie (Chemical Monthly), 148, 1983–1991.
Storbeck K H, Swart A C, Fox C L, Swart P. 2015. Cytochrome b5 modulates multiple reactions in steroidogenesis by diverse mechanisms. The Journal of Steroid Biochemistry and Molecular Biology, 151, 66–73.
Storbeck K H, Swart A C, Goosen P, Swart P. 2013. Cytochrome b5: Novel roles in steroidogenesis. Molecular and Cellular Endocrinology, 371, 87–99.
Strittmatter P. 1967. NADH-cytochrome b5 reductase. Methods in Enzymology, 10, 561–565.
Traylor M J, Baek J M, Richards K E, Fusetto R, Huang W, Josh P, Chen Z, Bollapragada P, O’Hair R A, Batterham P, Gillam E M. 2017. Recombinant expression and characterization of Lucilia cuprina CYP6G3: Activity and binding properties toward multiple pesticides. Insect Biochemistry and Molecular Biology, 90, 14–22.
Vergeres G, Waskell L. 1995. Cytochrome b5, its functions, structure and membrane topology. Biochimie, 77, 604–620.
Yang M, Zhang J, Zhu K Y, Xuan T, Liu X, Guo Y, Ma E. 2009. Mechanisms of organophosphate resistance in a field population of oriental migratory locust, Locusta migratoria manilensis (Meyen). Collaboration with the Entomological Society of America, 71, 3–15.
Zhang X, Dong J, Wu H, Zhang H, Zhang J, Ma E. 2019. Knockdown of cytochrome P450 CYP6 family genes increases susceptibility to carbamates and pyrethroids in the migratory locust, Locusta migratoria. Chemosphere, 223, 48–57.
Zhang X, Kang X, Wu H, Silver K, Zhang J, Ma E, Zhu K Y. 2018. Transcriptome-wide survey, gene expression profiling and exogenous chemical-induced transcriptional responses of cytochrome P450 superfamily genes in migratory locust (Locusta migratoria). Insect Biochemistry and Molecular Biology, 100, 66–77.
Zhang X, Wang J, Liu J, Li Y, Liu X, Wu H, Ma E, Zhang J. 2017. Knockdown of NADPH-cytochrome P450 reductase increases the susceptibility to carbaryl in the migratory locust, Locusta migratoria. Chemosphere, 188, 517–524.
Zhao X, Gou X, Qin Z, Li D, Wang Y, Ma E, Li S, Zhang J. 2017. Identification and expression of cuticular protein genes based on Locusta migratoria transcriptome. Scientific Reports, 7, 45462.
Zhu F, Sams S, Moural T, Haynes K F, Potter M F, Palli S R. 2012. RNA interference of nadph-cytochrome P450 reductase results in reduced insecticide resistance in the bed bug, Cimex lectularius. PLoS ONE, 7, e31037.
Zhu W, Yu R, Wu H, Zhang X, Liu Y, Zhu K Y, Zhang J, Ma E. 2016. Identification and characterization of two CYP9A genes associated with pyrethroid detoxification in Locusta migratoria. Pesticide Biochemistry and Physiology, 132, 65–71.
[1] LI Tian-pu, ZHANG Li-wen, LI Ya-qing, YOU Min-sheng, ZHAO Qian. Functional analysis of the orphan genes Tssor-3 and Tssor-4 in male Plutella xylostella[J]. >Journal of Integrative Agriculture, 2021, 20(7): 1880-1888.
[2] MENG Miao, YU Qi, WANG Qin, LIU Chun, LIU Zhao-yang, REN Chun-jiu, CUI Wei-zheng, LIU Qing-xin. BmApontic is involved in neurodevelopment in the silkworm Bombyx mori[J]. >Journal of Integrative Agriculture, 2020, 19(6): 1439-1446.
[3] MA Mei-qi, HE Wan-wan, XU Shi-jing, XU Le-tian, ZHANG Jiang.
RNA interference in Colorado potato beetle (Leptinotarsa decemlineata): A potential strategy for pest control
[J]. >Journal of Integrative Agriculture, 2020, 19(2): 428-427.
No Suggested Reading articles found!