Kinetic analysis of ribosome binding process onto mRNA using a quartz-crystal microbalance.

Translation initiation is the most dynamic and important step along a series of protein synthesis processes. In bacteria, it is generally accepted that the 70S ribosome initially dissociates into the 30S and 50S subunit, and then, the 30S ribosomal subunit binds to the Shine-Dargalno (SD) sequence of mRNA. We analyzed binding kinetics of 70S, 50S and 30S ribosomes to the SD sequence by using a mRNA-immobilized 27 MHz quartz-crystal microbalance (QCM). The 70S ribosome was found to bind strongly to the SD sequence as a ratio of 1:1 without dissociation to each subunit from the lateral side, as well as the 30S subunit. The binding constant for 70S increased in the presence of the initiator tRNA, which suggests that the SD and initiator codon of AUG could be also recognized precisely with 70S.