Abstract
Various types of mutants of precursor to the small subunit of RUBISCO (prSS) were designed and overexpressed in E. coli. The wild-type of Pisum sativum prSS has three Cystein residues, while these mutants have single Cystein residue at the various positions. These single-Cystein mutants (prSSC1) were modified with either the photoreactive crosslinker or the biotinylation reagent at the Cystein residue. The modifications didn't affect both the binding and the import activities of prSSC1. The protein close to one of the prSSC1 during binding step was crosslinked with prSSC1 via photoreactive crosslinker. Thus the identification of this crosslinked product is underway. On the other hand, one of the biotinylated prSSC1, incubated with avidin prior to the import assay, formed the protein translocation intermediate, in which prSSC1 was processed, but remained at the membrane. The characterization of the entire complex of this protein translocation intermediate is also underway.
