Reconstitution of Core Light-Harvesting Complexes of Photosynthetic Bacteria Using Chemically Synthesized Polypeptides. 1. Minimal Requirements for Subunit Formation

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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 β-polypeptides of the core light-harvesting complex (LH1) of Rhodobacter sphaeroides or Rhodospirillum rubrum. The native β-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 β-polypeptide of Rb. sphaeroides (sphβ16) 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 β-polypeptide of Rb. sphaeroides (sphβ31) or the equivalent 31 amino acids of the β-polypeptide of Rs. rubrum (rrβ31) were fully competent in forming a subunit-type complex and exhibited association constants for complex formation comparable to or exceeding those of the native β-polypeptides. The absorption and CD spectra of these subunit-type complexes were nearly identical to those of subunit complexes formed with native β-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 α-polypeptides to form a LH1-type complex. However, sphβ31 formed a LH1-type complex absorbing at 849 nm without an α-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.