Assembly of light-harvesting bacteriochlorophyll in a model transmembrane helix in its natural environment

The transmembrane, bacteriochlorophyll-binding region of a bacterial light-harvesting complex, (LH2-alpha from the photosynthetic bacterium Rhodobacter sphaeroides) was redesigned and overexpressed in a mutant of Rb. sphacroides lacking LH2. Bacteriochlorophyll served as internal probe for the fitness of this new region for the assembly and energy transfer function of the LH2 complex. The ability to absorb and transfer light energy is practically undisturbed by the exchange of the transmembrane segment, valine - 7 to threonine + 6, of LH2-alpha with a 14 residue Ala-Leu sequence. This stretch makes up the residues of the transmembrane helix that are in close contact (less than or equal to4.5 Angstrom) with the bacteriochlorophyll molecules that are coordinated through His of both the a and beta-subunits. In this Ala-Leu stretch, neither alpha-His0, which binds the bacteriochlorophyll, nor the adjacent alpha-Ile-1, were replaced. Novel LH2 complexes composed of LH2-a with a model transmembrane sequence and a normal LH2-beta are assembled in vivo into a complex, the biochemical and spectroscopic properties of which closely resemble the native one. In contrast, the additional insertion of four residues just outside the C-terminal end of the model transmembrane helix leads to complete loss of functional antenna complex. The results suggest that light energy can be harvested and transferred efficiently by bacteriochlorophyll molecules attached to only few key residues distributed over the polypeptide, while residues at the bacteriochlorophyll-helix interface seem to be largely dispensable for the functional assembly of this membrane protein complex. This novel antenna with a simplified transmembrane domain and a built-in probe for assembly and function provides a powerful model system for investigation of the factors that contribute to the assembly of chromophores in membrane embedded proteins. (C) 2002 Elsevier Science Ltd. All rights reserved.