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Expression of a bee venom phospholipase A2 from Apis cerana cerana in the baculovirus-insect cell

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Abstract

Bee venom phospholipase A2 (BvPLA2) is a lipolytic enzyme that catalyzes the hydrolysis of the sn-2 acyl bond of glycerophospholipids to liberate free fatty acids and lysophospholipids. In this work, a new BvPLA2 (AccPLA2) gene from the Chinese honeybee (Apis cerana cerana) venom glands was inserted into bacmid to construct a recombinant transfer vector. Tn-5B-4 (Tn) cells were transfected with the recombinant bacmid DNA for expression. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed a double band with molecular weights of 16 and 18 kDa. Products of hexahistidine AccPLA2 fusion protein accumulated up to 5.32% of the total cellular proteins. The AccPLA2 fusion protein was cross reactive with the anti-AmPLA2 (BvPLA2 of the European honeybee, Apis mellifera) polyclonal serum. The reaction resulted in a double glycosylation band, which agrees with the band generated by the native AmPLA2 in Western blot analysis. The PLA2 activity of the total extracted cellular protein in the hydrolyzing egg yolk is about 3.16 μmol/(min·mg). In summary, the recombinant AccPLA2 protein, a native BvPLA2-like structure with corresponding biological activities, can be glycosylated in Tn cells. These findings provided fundamental knowledge for potential genetic engineering to produce AccPLA2 in the pharmaceutical industry.

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Authors and Affiliations

  1. Department of Food Science and Nutrition, Zhejiang University, Hangzhou, 310029, China

    Li-rong Shen, Mei-hui Ding, Li-wen Zhang, Wei-guang Zhang & Duo Li

  2. Department of Agriculture and Natural Resources, Delaware State University, Dover, DE, 19901, USA

    Liang Liu

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  1. Li-rong Shen
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  2. Mei-hui Ding
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  4. Wei-guang Zhang
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  5. Liang Liu
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  6. Duo Li
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Correspondence to Li-rong Shen.

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Project (No. 2007AA10Z324) supported by the National High-Tech Research and Development Program (863) of China

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Shen, Lr., Ding, Mh., Zhang, Lw. et al. Expression of a bee venom phospholipase A2 from Apis cerana cerana in the baculovirus-insect cell. J. Zhejiang Univ. Sci. B 11, 342–349 (2010). https://doi.org/10.1631/jzus.B0900254

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  • DOI: https://doi.org/10.1631/jzus.B0900254

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