High level expression of soluble glycoproteins in the allantoic fluid of embryonated chicken eggs using a Sendai virus minigenome system

Soluble forms of human respiratory syncytial virus (HRSV) and human metapneumovirus (HMPV) fusion (F) proteins, devoid of their transmembrane and cytoplasmic domains, were produced in allantoic fluids using the Sendai minigenome system. The first step was rescuing in cell cultures Sendai virus minigenomes encoding the proteins of interest, with the help of wild type Sendai virus. The second step was propagating such recombinant defective viruses, together with the helper virus, in the allantoic cavity of chicken embryonated eggs, and passage to optimize protein production. When compared with the production of the same proteins in the culture supernatant of cells infected with vaccinia recombinants, the yield in the allantoic fluid was 5–10 fold higher. Mutant forms of these soluble proteins were easily constructed by site-directed mutagenesis and expressed in eggs using the same approach.The simplicity and economy of the Sendai minigenome system, together with the high yield achieved in the allantoic fluid of eggs, makes it an attractive method to express soluble glycoproteins aimed for structural studies.Over the past decades different expression systems have been developed for production of recombinant proteins. Each of these systems has strengths and weaknesses concerning yield, cost, speed, ease of manipulation and folding and post-translational modifications of the target proteins. E. coli is the simplest and most widely used organism for protein expression due to low cost and ease of use but it has serious limitations for expression of mammalian gene products, particularly glycoproteins [1]. Unmodified yeasts, as eukaryotes, are suitable for the production of proteins that do not require mammalian-type glycosylation [2]. However, cultured animal cells still remain the best system in which to produce mammalian glycoproteins, although they have complex nutritional requirements and are sensitive to viral and bacterial contamination [1].A repertoire of animal viru