The lysosomal enzyme acetyl CoA:alpha-glucosaminide N-acetyltransferase (GNAT) was shown to be an integral membrane protein requiring high concentrations of the detergent Triton X-100 for maximal solubilization. Using a concentration dependent Triton X-100 solubilization procedure and Concanavalin A-Sepharose affinity chromatography, GNAT was purified 50-fold with a yield of 45%. GNAT activity was separated from N-acetyltransferase activity toward glucosamine 6-phosphate, an alternative non-lysosomal pathway for glucosamine metabolism. GNAT was different from other lysosomal enzymes which bound to Concanavalin A-Sepharose in that both alpha-methylmannoside and Triton X-100 were required for elution of enzyme activity. GNAT activity, which bound to Concanavalin A-Sepharose, required at least one other component which did not bind for maximal expression of enzyme activity and for storage stability. Phospholipids and glycolipids, such as phosphatidylethanolamine, phosphatidylcholine, phosphatidylglycerol, sphingomyelin and gangliosides, and bovine serum albumin allowed expression of enzyme activity and storage stability similar to the component(s) which did not bind to Concanavalin A-Sepharose.
|Number of pages||9|
|Publication status||Published or Issued - May 1983|
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