Abstract
Plant-based expression systems have emerged as a competitive platform in the large-scale production of recombinant proteins. By adding a signal peptide, αAmy3sp, the desired recombinant proteins can be secreted outside transgenic rice cells, making them easy to harvest. In this work, to improve the secretion efficiency of recombinant proteins in rice expression systems, various signal peptides including αAmy3sp, CIN1sp, and 33KDsp have been fused to the N-terminus of green fluorescent protein (GFP) and introduced into rice cells to explore the efficiency of secretion of foreign proteins. 33KDsp had better efficiency than αAmy3sp and CIN1sp for the secretion of GFP from calli and suspension-cultured cells. 33KDsp was further applied for the secretion of mouse granulocyte-macrophage colony-stimulating factor (mGM-CSF) from transgenic rice suspension-cultured cells; approximately 76%–92% of total rice-derived mGM-CSF (rmGM-CSF) was detected in the culture medium. The rmGM-CSF was bioactive and could stimulate the proliferation of a murine myeloblastic leukemia cell line, NSF-60. The extracellular yield of rmGM-CSF reached 31.7 mg/L. Our study indicates that 33KDsp is better at promoting the secretion of recombinant proteins in rice suspension-cultured cell systems than the commonly used αAmy3sp.
Highlights
Plant-based protein expression systems have been successfully used to produce several recombinant proteins [1,2,3,4]
The production of recombinant proteins in cell cultured medium have been conducted in several recombinant protein expression systems
Recombinant proteins that require post-translational modification in order to retain bioactivity or stability can be modified through a series of enzymes located in the endoplasmic reticulum (ER) and the Golgi apparatus before secretion
Summary
Plant-based protein expression systems have been successfully used to produce several recombinant proteins [1,2,3,4]. A signal peptide derived from the resident ER protein calreticulin was applied in a plant hair root system [17,18]; in addition, an α-amylase signal peptide was used in a rice suspension-cultured cell system [8]. It was reported that the recombinant proteins were not secreted efficiently into extracellular space, and a large proportion of recombinant proteins still remained within the cells [19]. This might have been due to the low activity of the applied signal peptide. The detailed mechanism is not clear yet for the correlation between signal peptide and protein secretion, one proposed model is that signal peptide plays an important role to lead recombinant protein into the lumen of the ER [24,25,26], which is one of limiting steps of protein secretion in a default pathway
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