Isolation and characterization of an organic solvent soluble polypeptide component from photoreceptor complexes of Rhodospirillum rubrum

Abstract

An organic solvent soluble polypeptide has been isolated from photoreceptor complexes and chromatophores of Rhodospirillum rubrum. After extraction of the protein from lyophilized samples with 1:1 chloroform-methanol, it was purified by column chromatography. Its isoelectric point determined by isoelectric focusing was 7.10. When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the purified polypeptide ran as a single band of an apparent molecular weight of 12 000. However, according to amino acid analysis, the minimal molecular weight based on one histidine residue per polypeptide is 19 000. The polypeptide contains no cysteine and no tyrosine. Amino acid analysis indicated that three methionines were present per histidine residue and cyanogen bromide cleavage gave four smaller peptides which were isolated by two-dimensional electrophoresis and chromatography. Spectroscopic analysis indicated the presence of three tryptophan residues per histidine and N-bromosuccinamide cleavage also gave four smaller peptides which could be isolated by two-dimensional electrophoresis and chromatography. The C-terminal amino acid was shown to be glycine by two methods, while the N-terminal amino acid appears to be blocked. The organic solvent soluble polypeptide accounts for approximately 50% of the chromatophore protein and seems to bind the antenna bacteriochlorophyll and carotenoid molecules. Using this procedure, organic solvent soluble polypeptides were isolated from several photosynthetic bacteria and were found to have substantially different amino acid contents.

Isolation and Characterization of the Polypeptide Components from Light‐Harvesting Pigment‐Protein Complex B800–850 of Rhodopseudomonas capsulata

“…Unlike the organic solvent-soluble polypeptide isolated from R. ruhrum chromatohores by Tonn and coworkers [20] and Cuendet and Zuber [22], the N-terminal a-amino groups of the two smaller polypeptides from the B800-850 complex are not blocked. Fraker and Kaplan [23,24] have partially characterized a low molecular-weight protein isolated from R. .s/dzaeroides chromatophores by extraction with 2-chloroethanol.…”

“…1). Initially the chloroform!methanol system employed by Tonn and coworkers [20] was tried; however, in the absence of salt the three polypeptides were eluted as one band just ahead of the pigments (data not shown here). Sandermann and Strominger [21] obtained a fraction enriched in Cssisoprenoid alcohol phosphokinase after gel filtration on Sephadex LH-60 in a mixture of butanol/methanol containing 1 M ammonium acetate.…”

“…The polypeptide of the light-harvesting bacteriochlorophyll-protein of Rliodospirillum ruhrum has been recently isolated by gel filtration in organic solvent [20, 221. Organic solvent soluble polypeptides have been also reported and, to various extents, characterized in [20] have reported isolating an organic solvent-soluble polypeptide from wild-type R. capsulatu; unfortunately, characterization data from this preparation were no1 available. In general, the amino acid contents of these proteins are typical of membrane constituents.…”

The complete amino acid sequence of the single light harvesting protein from chromatophores of Rhodospirillium rubrum G‐9+

“…Based on the amino acid analysis after 240 h hydrolysis, LHP has Mr -6000. The much lower number of 73 amino acid residues/l His residue after 24 h ( fig.1) compared to [ 1,2] is apparently the result of the improved chromatographic purification of LHP on Sephadex LH-60 in C/M/NHGOAc. LHPl Deblocking conditions for LHP were improved by taking 5% HCl in C/M (1: 1, v/v) instead of 1 N aqueous HCl [6].…”

“…Pigment-free preparations of the single light-harvesting protein from the purple photosynthetic bacterium Rhodospirillum rubrum have been reported both from the antenna bacteriochlorophyll-protein complex and directly from chromatophores by chloroform/methanol extraction [ 1,2]. Molecular mass determinations yielded values of 9000-l 9 000 [ l-4 1.…”