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Met129 is required for lyLMP-1 translation. The lyLMP-1 ORF was expressed in vitro from the T7 promoter in pcDNA3-based expression plasmids encoding Met129 (+ATG) or Ala129 (-ATG). Translated products were resolved by SDS-PAGE and analyzed by Western blotting using affinity-purified anti-LMP-1 rabbit antiserum recognizing LMP-1's carboxy terminus (14) followed by anti-rabbit-horseradish peroxidase-conjugated secondary antibody. lyLMP-1 was visualized by ECL (Amersham).
