A simple reproducible cell-free system for measuring brain protein synthesis.
A simple, rapid, sensitive, and reproducible cell-free assay system for studying brain protein synthesis is described. This system uses small amounts of brain postmitochondrial supernatant, making it a convenient screening test when only small amounts of tissue are available. It showed over 95% dependence on Mg2+ and on an energy source. Optimal incorporation occurred under the following conditions: Mg2+ 3 mM; ATP, 0.6 mM; GTP, 0.6 mM; high K+, greater than or equal to 25 mM; Low Na+, less than or equal to 15 mM; pH 7.1-7.5. The rate of amino acid incorporation did not vary with leucine concentrations in vitro up to 1 mM, which obviated the need to measure endogenous leucine concentrations.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002474:Cell-Free System; D006160:Guanosine Triphosphate / Q000494:pharmacology; D007930:Leucine / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008861:Microsomes / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000096:biosynthesis*; D051381:Rats; D012964:Sodium / Q000494:pharmacology; D014316:Tritium
D000596:Amino Acids; D009419:Nerve Tissue Proteins; D014316:Tritium; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D012964:Sodium; D007930:Leucine; D008274:Magnesium