Protein synthesis and competitive ESR binding studies with E. coli ribosomes and spin-labeled polynucleotides

Enzymatically prepared spin labeled copolymers of (U)n were tested for their ability to direct polyphenylalanine synthesis in vitro using E. coli B enzymes and ribosomes. Spin labeling of the C5 position using (RUGT,U)n (1:100) or (RUTT,U)n (1:100) did not alter the amount of polyphenylalanine formed in comparison to (U)n. In contrast, the C4 spin labeled copolymer (ls4U,U)n (1:100) reduced phenylalanine incorporation by 70-75% of the (U)n control levels. ESR monitoring of competitive ribosome binding to equimolar mixtures of polynucleotides was demonstrated with the macromolecular probe (DUTT,dT)n (1:100), the DNA analogue of (RUTT,U)n. The ESR competition approach showed that the affinity of the ribosomes was essentially the same for (dT)n, (A,U,G)n, and (A,U,G)n + tRNArmet.