Photoaffinity Labeling of the Rubisco Active-Site and Small Subunit with 8-Azidoadenosine 5′-Triphosphate

Abstract

The 8-azidopurine analogs have been used as probes for identifying nucleotide-binding proteins in many biological systems (1). Irradiation with UV light converts the azido moiety to a reactive species that can bind covalently to residues in or near the nucleotide site. In the present study, photoaffinity labeling was used to probe for possible binding sites on the subunits of Rubisco. The results presented herein provide evidence for distinct binding sites on the Rubisco subunits by showing that [³²P]8-N₃ATP labels four active site peptides of the large subunit (LSu) and two peptides from conserved regions of the small subunit (SSu).