Design of a minimal polypeptide unit for bacteriochlorophyll binding and self-assembly based on photosynthetic bacterial light-harvesting proteins.
We introduce LH1beta24, a minimal 24 amino acid polypeptide that binds and assembles bacteriochlorophylls (BChls) in micelles of octyl beta-glucoside (OG) into complexes with spectral properties that resemble those of B820, a universal intermediate in the assembly of native purple bacterial light-harvesting complexes (LHs). LH1beta24 was designed by a survey of sequences and crystal structures of bacterial LH proteins from different organisms combined with currently available information from in vitro reconstitution studies and genetically modified LHs in vivo. We took as a template for the design sphbeta31, a truncated 31 amino acid analogue of the native beta-apoprotein from the core LH complex of Rhodobacter sphaeroides. This peptide self-assembles with BChls to form B820 and, upon cooling and lowering OG concentration, forms red-shifted B850 spectral species that are considered analogous to native LH complexes. We find that LH1beta24 self-assembles with BChl in OG to form homodimeric B820-type subunits comprising two LH1beta24 and two BChl molecules per subunit. We demonstrate, by modeling the structure using the highly homologous structure of LH2 from Rhodospirillum molischianum, that it has the minimal size for BChl binding. Additionally, we have compared the self-assembly of sphbeta31 and LH1beta24 with BChls and discovered that the association enthalpies and entropies of both species are similar to those measured for native LH1 from Rhodospirillum rubrum. However, sphbeta31 readily aggregates into intermediate higher oligomeric species and further to form B850 species; moreover, the assembly process of these oligomers is not reversible, and they are apparently large nonspecific BChl-peptide coaggregates rather than well-defined nativelike LH complexes. Similar aggregates were observed during LH1beta24 assembly, but these were formed less readily and required lower temperatures than sphbeta31. In view of these results, we reevaluate previous in vitro reconstitution studies and propose alternative templates for new designs.
Noy
Dror
D
Johnson Research Foundation, Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA. dror.noy@weizmann.ac.il
Dutton
P Leslie
PL
D000595:Amino Acid Sequence; D001429:Bacteriochlorophylls / Q000378:metabolism*; D002498:Centrifugation; D002942:Circular Dichroism; D045342:Light-Harvesting Protein Complexes / Q000378:metabolism*; D008969:Molecular Sequence Data; D045331:Photosystem I Protein Complex / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods; D016415:Sequence Alignment; D013053:Spectrophotometry; D013816:Thermodynamics
D001429:Bacteriochlorophylls; D045342:Light-Harvesting Protein Complexes; D045331:Photosystem I Protein Complex