Regeneration of native bacteriorhodopsin structure following acetylation of epsilon-amino groups of Lys-30, -40, and -41.
The chymotryptic fragment of bacteriorhodopsin, C-2 (residues 1-71), has been acetylated completely at its three lysines (residues 30, 40, and 41) by treatment with acetic anhydride. The triacetylated C-2 fragment is able to reassociate with fragment C-1 (residues 72-248) and the complex binds all-trans-retinal to form a native bacteriorhodopsin-like chromophore, which is essentially identical with that formed from fragments C-2 and C-1. Further, the kinetics and pH dependence of chromophore regeneration and the proton pumping of the reconstituted triacetylated C-2 and C-1 complex are indistinguishable from that of the unmodified C-2 and C-1 complex. However, the extent of regeneration of the chromophore from triacetylated C-2 and C-1 is less than that from fragments C-2 and C-1, suggesting that the acetylated C-2 fragment is less stable than unacetylated C-2 in the reconstitution medium. We conclude that the amino groups in Lys-30, -40, and -41 do not contribute to the stabilization of the folded bacteriorhodopsin structure and are not required for proton translocation.
D000089:Acetic Anhydrides; D000107:Acetylation; D001436:Bacteriorhodopsins / Q000378:metabolism*; D002338:Carotenoids / Q000378:metabolism*; D002918:Chymotrypsin / Q000378:metabolism; D002942:Circular Dichroism; D006217:Halobacterium / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008239:Lysine; D010446:Peptide Fragments / Q000378:metabolism; D011487:Protein Conformation; D011522:Protons; D012172:Retinaldehyde / Q000378:metabolism; D013053:Spectrophotometry
D000089:Acetic Anhydrides; D010446:Peptide Fragments; D011522:Protons; C031800:acetic anhydride; D002338:Carotenoids; D001436:Bacteriorhodopsins; D002918:Chymotrypsin; D008239:Lysine; D012172:Retinaldehyde