Measurement of Ribosomal Accuracy and Proofreading in E. coli Burst Systems

Abstract

The programmed Escherichia coli ribosome chooses one tRNA isoacceptor from approx 40 competing aminoacyl-tRNAs (1). If an aminoacyl-tRNA with an anticodon nonmatching to the codon is accepted by the ribosome, an erroneous amino acid may be incorporated in the nascent polypeptide, with a missense error as a result. Missense errors in E. coli vary considerably depending on codons and contexts (2). A global average missense frequency has been estimated to be in the range of 3×10⁻⁴ per codon for wild-type ribosomes (3–6). With this error rate, approx 10% of proteins with an average length of 400 amino acids will contain a single missense error, whereas a ribosome, containing about 10,000 amino acids, will have three amino acid substitutions (7). There are ribosome variants with considerably lower (8) as well as with significantly higher (9) missense error levels than wild-type ribosomes. Bacteria with hyperaccurate as well as with error-prone ribosomes grow slower than otherwise isogenic wild-type cells. This suggests that the error level of wild-type ribosomes has evolved to maximize the bacterial growth rate (7): The maximum corresponds to the “best” compromise between the vices of too high accuracy and too many errors in the cell’s proteins. On one hand hyperaccurate ribosomes have impaired efficiency in their interaction with cognate ternary complexes and aa-tRNAs (10), and this tends to reduce the growth rate. Error-prone ribosomes, on the other, produce proteins (enzymes) containing one or several erroneous amino acids, and this reduces their kinetic efficiency and thereby the growth rate of the bacterial population (7).