Hyaluronate is synthesized at plasma membranes.
The hybrid cell B6 line, which synthesizes large amounts of hyaluronate as the predominant glycosaminoglycan, was grown in the presence of [3H]glucosamine. The [3H]hyaluronate has a high molecular weight and was excluded by Sephacryl S-1000. After disruption of the cells the [3H]hyaluronate could further be elongated by incubation with UDP-GlcNAc and UDP-[14C]GlcA, yielding a hybrid molecule of hyaluronate labelled with [3H]GlcNAc and [14C]GlcA. Treatment of the cells with hyaluronidase before disruption eliminated the large [3H]hyaluronate and elongation of nascent chains in vitro commenced from low-molecular-weight chains. Thus nascent hyaluronate chains were degraded extracellularly by hyaluronidase and were therefore synthesized at the inner side of plasma membranes and extruded to the cell surface.