Kinetics of translation of γB crystallin and its circularly permutated variant in an in vitro cell‐free system: possible relations to codon distribution and protein folding

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Abstract
Analysis of nascent γB-crystallin peptides accumulating during in vitro translation in a rabbit reticulocyte lysate cell-free system was carried out. As a consequence of the irregular distribution of rare codons along the polypeptide chain of γB-crystallin, translation of the two-domain protein is a non-uniform process characterized by specific pauses. One of the major delays occurs during the translation of the connecting peptide between the domains. Comparing the kinetics of translation of natural γB-crystallin and its circularly permutated variant (with the order of the N- and C-terminal domains exchanged) reveals that the natural N-terminal domain is translated faster than the C-terminal one. Since the N-terminal domain in natural γB-crystallin is known to be more stable and to fold faster than the C-terminal one [E.-M. Mayr et al. (1994) J. Mol. Biol. 235, 84–88], the present data suggest that the translation rates are optimized to tune the synthesis and folding of the nascent polypeptide chain. In this connection, the pause in the linker region between the domains provides a delay allowing the correct folding of the N-terminal domain and its subsequent assistance in the stabilization of the C-terminal one