High-density tissue-like cultivation of JAR choriocarcinoma cells for the in vitro production of human xylosyltransferase

Abstract

Human xylosyltransferase is the chain-initiating enzyme involved in the biosynthesis of glycosaminoglycans. Large amounts of xylosyltransferase are required to study the biochemical properties of the native enzyme. To achieve this goal a scale-up of animal cell culture systems was inevitable due to the small amounts of the enzyme present in tissues, e.g. only 0.5 μg XT can be obtained from a chick embryo. JAR choriocarcinoma cells cultured with 10% fetal calf serum were found to secrete xylosyltransferase with relatively high activities (1.10 mU l −1). To reduce contaminating proteins JAR cells were adapted to serum-free conditions. Xylosyltransferase activities up to 0.22 mU l −1 were determined in the harvested cell culture supernatant. Scaling-up of JAR cell culture in the hybrid hollow fiber bioreactor Tecnomouse resulted in the production of 15.8 mU or 270 μg XT in 0.5 l of XT-enriched cell culture supernatant using 57 l of serum-free cell culture medium. The XT activity per ml harvest solution was 200–280-fold higher in this cell culture supernatant than in cell culture flasks. In addition, the specific XT activity of the bioreactor product was 6 μU mg −1 of total protein, which is 2-fold higher than that obtained under static culture conditions. This study clearly demonstrates the successful high-density, tissue-like cultivation of JAR choriocarcinoma cells in a hollow fiber bioreactor resulting in an effective production of native human xylosyltransferase.