Importance of the carboxyl terminus of human thyroid peroxidase in the efficient expression of the protein in eukaryotic cells.

摘要: Abstract Carboxyl terminal truncation of membrane-associated human thyroid peroxidase (hTPO), with the elimination its single membrane-spanning and short intracytoplasmic regions, generates a soluble, secreted, enzymatically active protein (amino acids 1–848). In order to determine effects further carboxyl deletions on expression hTPO, Chinese hamster ovary cells were stably transfected plasmids constructed express amino 1–771, 1–636, 1–539 1–382 933 acid TPO protein, respectively. Unlike hTPO1–848, more severely truncated mutant proteins could not be detected in conditioned media by polyclonal anti-TPO antibodies. Using detergent-solubilized microsomal from these cells, very low levels hTPO1–771 (approximately 90 kDa), but extensive deletion mutations, Confirmation loss efficient hTPO was obtained using anti-hTPO monoclonal antibody an epitope near terminus which recognizes only denatured protein. The mRNA for all mutants stably-transfected cells. summary, present study indicates that largely intact extracellular portion is required eukaryotic