Figure 2.

Immunoprecipitation of Menkes ATPase from HeLa cell lysates. Following a 1-h pulse-label with [³⁵S]methionine and [³⁵S]cysteine, cells were lysed and immunoprecipitated with preimmune sera (lane 1), Menkes antibody (lanes 2, 4, and 6–9), or Menkes antibody preincubated with Menkes fusion protein (lanes 3 and 5). For some experiments cells were pretreated with tunicamycin (4 μg/ml) for 2 h prior to immunoprecipitation (lanes 4, 5, and 9). Immunoprecipitated Menkes protein was digested with endo H or endo F (lanes 7 and 8) as described. Samples were electrophoresed on SDS/5-12% polyacrylamide gradient gels and subjected to fluorography.