Following formation of a peptide bond, the P site of the ribosome contains an uncharged tRNA, and the A site contains a tRNA linked to the growing peptide chain. Translocation is the process of recocking the elongation mechanism. The uncharged tRNA in the P site is moved to the exit, or E site, messenger translocates three bases toward the P site, thereby moving the tRNA with the peptide chain into the P site.
The translocation process itself requires hydrolysis of a GTP molecule that has been carried to the ribosome by the EF-G or G factor. Since the elongation factors are used once for each amino acid added, a large number of molecules of each must be present in the cell to support protein synthesis. It is also logical that their level should parallel the level of ribosomes, and, indeed, as growth rate varies, their levels do keep pace with the levels of ribosomes. With the entry of a charged tRNA into the A site of the ribosome, the uncharged tRNA in the E site is released.
At some time during the growth of the peptide chains, the N-terminal amino acid is modified. Approximately 40% of the proteins isolated from E. coli are found to begin with methionine, but since all initiate with N-formyl methionine, the remaining 60% must lose at least the N-terminal methionine. Similarly, the 40% of the proteins that do begin with methionine all lack the formyl group. Thus the formyl group must be removed after protein synthesis has initiated. Examination of nas-cent polypeptide chains on ribosomes shows that the formyl group is missing if they are larger than about 30 amino acids, and therefore the deformylase could well be a part of the ribosome. It could act when the growing peptide chain is long enough to reach the enzyme.
The deformylase is a very labile enzyme that is exceedingly sensitive to sulfhydryl reagents. Since many other enzymes isolated from the same cells require the same sulfhydryl reagents for stability, it may be that the deformylase is normally bound to some structure that contrib-utes to its stability, and when it is isolated from extracts and partially purified, it is particularly labile in its unnatural environment.
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