Synthesis of functional aldolase tetramers in a heterologous cell-free system.

Abstract

The present report describes the complete synthesis of a functional oligomeric enzyme in a heterologous cell-free system. Polysomal RNA from chicken skeletal muscle was used to direct the production of functional aldolase tetramers in wheat germ extracts. The aldolase product was (a) specifically precipitated with monospecific antibodies raised against pure muscle aldolase, (b) had the same subunit molecular weight (40,000) as that of native aldolase (as determined by acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate), (c) presumably contained a functional active site since it co-purified with authentic muscle aldolase upon substrate elution from phosphocellulose, and (d) had associated into tetrameric units (Mr=160,000) as shown by centrifugation in sucrose gradients. The present work suggests that, within the cell, post-translational processing of aldolase polypeptide chains is not involved in the formation of functional aldolase tetramers.