THE ROLE OF THE LEADER SEQUENCE CODING REGION IN EXPRESSION AND ASSEMBLY OF BACTERIORHODOPSIN

Bacterio-opsin is made as a precursor in Halobacterium halobium, which has 13 additional residues at the amino terminus. The codons for thes e residues have been proposed to form a hairpin structure in the mRNA and play a role in ribosome binding; the leader peptide sequence also has been proposed to have a role in membrane insertion of bacteriorhod opsin (BR). We have made mutations in the bop gene region coding for t he leader sequence and expressed the mutant genes in an H. halobium mu tant lacking wild-type BR. The leader sequence coding region was found to be important for the stability of the mRNA and for its efficient t ranslation. Single base substitutions in this region that did not affe ct the amino acid sequence caused significant reductions in protein ex pression. Deletion of the leader region resulted in unstable mRNA and almost no BR production. Introduction of a new ribosome-binding sequen ce within the coding region of the mature protein restored mRNA stabil ity and some protein expression, Protein made without the leader pepti de was properly assembled in the membrane.