Plastocyanin of Arabidopsis thaliana; isolation and characterization of the gene and chloroplast import of the precursor protein.

A genomic clone encoding the plastocyanin precursor was isolated from an Arabidopsis thaliana lambda EMBL3 library, with the help of a heterologous hybridization probe. The nucleotide (nt) sequence encoding the 171-amino acid precursor protein and 650 bp of the 5'-flanking region were determined. S1 nuclease mapping showed the Arabidopsis coding region to be uninterrupted and the transcript to possess an untranslated leader of approx. 30 nt. The 5' region of the gene contains a 25-bp direct repeat at a distance of 300 bp upstream from the ATG start codon. Southern analysis of several genomic digests shows the presence of a single copy of the plastocyanin gene in the Arabidopsis genome. In vitro synthesized pre-plastocyanin was used in import experiments with isolated pea chloroplasts. Plastocyanin was correctly directed to the thylakoid lumen and processed to the mature size. A clear single processing intermediate, as was found with the import of Silene pratensis pre-plastocyanin, seems to be absent.