Purification of gamma-enolase messenger ribonucleic acid from rat brain by an immunoadsorption method.
Messenger ribonucleic acid (mRNA) coding for the brain-specific protein gamma-enolase was isolated by an immunopurification procedure. Rat brain polysomes including nascent polypeptide chains were reacted with specific gamma-enolase antibody. The polysome-antibody complexes were subsequently adsorbed to protein A-Sepharose. After extensive washing, RNA was eluted and applied to an oligo(dT)-cellulose column. Purified mRNA was translated in vitro in a mRNA-dependent rabbit reticulocyte lysate system. The synthesized product was identical to gamma-enolase synthesized by free polysomes from rat brain. Immunoisolated gamma-enolase mRNA was enriched 380-fold compared to total mRNA extracted from free polysomes. This result indicates that low-abundance mRNAs may conveniently be isolated from brain tissue by immunoadsorption of polysomes.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D001923:Brain Chemistry; D007163:Immunosorbent Techniques; D010751:Phosphopyruvate Hydratase / Q000096:biosynthesis / Q000235:genetics*; D011132:Polyribosomes; D012333:RNA, Messenger / Q000302:isolation & purification*; D051381:Rats; D011919:Rats, Inbred Strains
D012333:RNA, Messenger; D010751:Phosphopyruvate Hydratase