Purification and sequencing of cyanogen bromide fragments from ribulosebisphosphate carboxylase/oxygenase from Rhodospirillum rubrum

As a part of the goal to determine the total sequence of Rhodospirillum rubrum ribulosebisphosphate carboxylase/oxygenase, the cyanogen bromide fragments were fractionated and sequenced (or partially sequenced). Twelve of the anticipated 14 peptides were obtained in highly purified form. The other two peptides were located, respectively, within a trytophanyl cleavage product (which overlapped with four CNBr fragments) and within an active-site peptide characterized earlier (which overlapped with three CNBr fragments). These overlaps coupled with amino and carboxyl terminal sequence information of the intact subunit and the availability of the sequence of the corresponding enzyme from higher plants permitted alignment of all fragments. Eight CNBr peptides were sequenced completely; four of the CNBr peptides consisted of more than 80 residues and were only partially sequenced as permitted by direct Edman degradation. Of the approximate 475 residues per subunit, 339 were placed in sequence. The lack of extensive conservation of primary structure between R. rubrum and higher plant carboxylases permits the tentative identifications of those regions likely to be functionally important.