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Enhanced expression of the human CD14 protein in tobacco using a 22-kDa alpha-zein signal peptide

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Abstract

The human CD14, a high affinity receptor for lipopolysaccharides (LPS), is involved in the innate immunity system and the inflammatory response. There is increasing interest in using recombinant approaches to produce purified CD14 protein for therapeutic uses. Plants provide ideal expression systems for the production of recombinant proteins, but the levels of expression of recombinant proteins produced in planta are still not high. To improve expression levels of CD14 the 22-kDa alpha-zein signal peptide (ZSP) from maize was fused to the human CD14 cDNA so that recombinant CD14 could stably accumulate in plant cells. The human CD14 gene and the modified human CD14 cDNA with the 22-kDa ZSP were respectively transformed into tobacco to produce transgenic plants. Western blot analysis confirmed human CD14 accumulation in the transgenic tobacco. The concentration of the recombinant protein in the tobacco leaves was measured by ELISA, and the results suggested that fusion with the 22-kDa alpha-ZSP effectively increased the accumulation of the recombinant protein (rCD14). The concentration of rCD14 in some of the transgenic lines was 19.54 μg g−1 tobacco leaf (fw), which was about 0.6 % of the total soluble protein. The rCD14 protein showed natural LPS-binding bioactivity by using U937 cells mensuration. Our results suggested that the maize 22-kDa alpha-zein signal peptide could be used to increase the accumulation of recombinant protein in tobacco leaves so that proteins can be produced in abundant biomass.

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Abbreviations

als :

Acetolactate synthase gene

CaMV:

Cauliflower mosaic virus

epsp :

5-Enolpyrul-shikimate-3-phosphate synthase gene

fw:

Fresh weight

GPI:

Glycosylphosphatidylinositol

LPS:

Lipopolysaccharide

mCD14:

Membrane-associated CD14

MW:

Molecular weight

PSV:

Protein storage vacuole

rCD14:

Recombinant CD14

sCD14:

Soluble CD14

WT:

Wild type

ZSP:

Zein signal peptide

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Acknowledgments

We thank Dr. Roberta Greenwood (Shandong University, China) for her help in editing this manuscript. This research was supported by National Basic Research Program of China (973 Program, 2009CB118400) and Natural Science Foundation of China (no. 30771127).

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Authors and Affiliations

  1. School of Life Science, Shandong University, 27 Shanda South Road, Jinan, 250100, People’s Republic of China

    Xiuxia Liu, Li Sun, Aifang Yang & Juren Zhang

  2. Institute of Basic Medicine, Shandong Academy of Medical Science, 18877 Jingshi Road, Jinan, 250062, People’s Republic of China

    Cuiling Li

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  1. Xiuxia Liu
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  2. Li Sun
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  3. Cuiling Li
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  4. Aifang Yang
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  5. Juren Zhang
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Correspondence to Juren Zhang.

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Liu, X., Sun, L., Li, C. et al. Enhanced expression of the human CD14 protein in tobacco using a 22-kDa alpha-zein signal peptide. Plant Cell Tiss Organ Cult 112, 9–18 (2013). https://doi.org/10.1007/s11240-012-0206-x

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  • DOI: https://doi.org/10.1007/s11240-012-0206-x

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