title: Reconstitution of core light-harvesting complexes of photosynthetic bacteria using chemically synthesized polypeptides. 1. Minimal requirements for subunit formation.
abstract: Described are the chemical synthesis, isolation and characterization of each of three polypeptides whose amino acid sequences reproduce portions of the amino acid sequence of the beta-polypeptides of the core light-harvesting complex (LH1) of Rhodobacter sphaeroides or Rhodospirillum rubrum. The native beta-polypeptides of LH1 of these organisms contain 48 and 54 amino acids, respectively. The smallest synthetic polypeptide had an amino acid sequence identical to that of the last 16 amino acids of the beta-polypeptide of Rb. sphaeroides (sph beta 16) but failed to form either a subunit- or LH1-type complex under reconstitution conditions. Also, this polypeptide, lengthened on the N terminus by adding the sequence Lys-Ile-Ser-Lys to enhance solubility, failed to form a subunit- or LH1-type complex. In contrast, polypeptides containing either the 31 amino acids at the C terminus of the beta-polypeptide of Rb. sphaeroides (sph beta 31) or the equivalent 31 amino acids of the beta-polypeptide of Rs. rubrum (rr beta 31) were fully competent in forming a subunit-type complex and exhibited association constants for complex formation comparable to or exceeding those of the native beta-polypeptides. The absorption and CD spectra of these subunit-type complexes were nearly identical to those of subunit complexes formed with native beta-polypeptides. It may be concluded that all structural features required to make the subunit complex are present in the well-defined, chemically synthesized polypeptides. Neither polypeptide appeared to interact with the native alpha-polypeptides to form a LH1-type complex. However, sph beta 31 formed a LH1-type complex absorbing at 849 nm without an alpha-polypeptide. Although chemical syntheses of polypeptides of this size are common, the purification of membrane-spanning segments is much more challenging because the polypeptides lack solubility in water. The chemical syntheses reported here represent the first such syntheses of membrane-spanning polypeptides which display native activity upon reconstitution.
authors.affiliation: Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, Illinois 60208-3500, USA.
authors.forename: K A
authors.initials: KA
authors.lastname: Meadows
authors.forename: P S
authors.initials: PS
authors.lastname: Parkes-Loach
authors.forename: J W
authors.initials: JW
authors.lastname: Kehoe
authors.forename: P A
authors.initials: PA
authors.lastname: Loach
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D002942:Circular Dichroism; D045342:Light-Harvesting Protein Complexes; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics; D045322:Photosynthetic Reaction Center Complex Proteins / Q000138:chemical synthesis* / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation; D012242:Rhodobacter sphaeroides / Q000737:chemistry / Q000235:genetics; D012247:Rhodospirillum rubrum / Q000737:chemistry / Q000235:genetics; D013045:Species Specificity; D013053:Spectrophotometry
chemical_list: D001426:Bacterial Proteins; D045342:Light-Harvesting Protein Complexes; D010446:Peptide Fragments; D045322:Photosynthetic Reaction Center Complex Proteins; C094744:light-harvesting complex 1, Rhodospirillum