Physical mapping of genes for chloroplast DNA encoded subunit polypeptides of the ATPsynthase complex from Petunia hybrida

Abstract

Escherichia coli minicells harbouring the cloned restriction fragment Sall S9 from P. hybrida chloroplast DNA synthesize the beta and epsilon polypeptide subunits of the CF₁ component of the chloroplast ATPsynthase complex. The polypeptides were identified by molecular weight determination and immunoprecipitation. The position of the atpB and the atpE gene, encoding respectively the beta and epsilon subunit, on the Sall S9 fragment was determined in more detail by studying polypeptide synthesis directed by subclones of the S9 fragment in E. coli minicells. The atpB and atpE genes are located close to the rbcL gene, the distance between the rbcL gene and atpB gene being approximately 770 bp. Analysis of the expression of subclones of the S9 fragment in E. coli minicells also revealed that the atpE gene can be transcribed and translated independently of the expression of the atpB gene. The location of the genes coding for the alpha subunit (atpA gene) and the proteolipid subunit III of CF₀ (atpH) of the ATPsynthase complex on the physical map of P. hybrida cpDNA was determined by hybridization of restriction enzyme digests of petunia cpDNA with cloned cpDNA fragments from Spirodela and wheat, containing internal parts of respectively the atpA and the atpH gene. The two genes map close together within a region of 5.2 kbp on the physical map of P. hybrida cpDNA. The distance between the atpA gene and the atpB and atpE genes is approximately 42 kbp.