ENGINEERING AND EXPRESSION OF A SECRETED MURINE TCR WITH REDUCED N-LINKED GLYCOSYLATION

Structural studies of TCR-alpha beta heterodimers would be greatly aid ed by the ability to produce nonchimeric, secreted material with less carbohydrate heterogeneity. Here, we report the engineering and expres sion of variants of the murine TCR 2B4 in which many of the potential N-linked glycosylation sites were eliminated. Specific truncations pro ximal to the transmembrane region were also introduced that result in a secreted heterodimer. Although elimination of N-linked oligosacchari de on the beta-chain does not significantly affect the expression leve ls of 2B4 heterodimers, ablation of N-linked oligosaccharide on the al pha-chain results in a measurable reduction in expression levels of me mbrane-associated molecules. Secreted forms of 2B4 heterodimers in whi ch the N-linked glycosylation of the beta-chain has been eliminated ca n be expressed. The secreted receptor is shown by a variety of Ab dete rminants to be indistinguishable from native material.