Ribosome nascent chain complexes of the chloroplast-encoded cytochrome b6 thylakoid membrane protein interact with cpSRP54 but not with cpSecY

We analysed the interplay between the cpSecY, cpSRP54 and the chloroplast-encoded cytochrome b(6) via isolation of chloroplast ribosome nascent chain complexes and the use of cross-linking factors, antibodies and mass spectroscopy analyses. We showed that the cytochrome b(6) nascent polypeptide complex is tightly associated with ribosomes and that the translation of cytochrome b(6) was discontinuous. The causes of ribosome pausing and the functional significance of this phenomenon may be related to proper protein folding, insertion into thylakoid membranes and the association of cofactors during this process. It was also found that cpSecY was not in the vicinity of cytochrome b(6) intermediates during the elongation process and does not act with mature cytochrome b(6) after translation. Using the approach of cross-linking during elongation of the cytochrome b(6) protein, we showed that cpSRP54 interacts strongly with the elongating nascent chain.