Major Proteins of the Amyloplast of Agar and Soil - Grown Potato Tubers

In potato (Solanum tuberosum) tuber starch is synthesized and stored in amyloplasts. Amyloplasts were prepared from in vitro or agar-grown micro tubers and from soil-grown mini tubers and subjected to proteome analysis. The quality of amyloplasts was assessed by comparing amyloplast fractions and total tuber extracts by SDS-PAGE and the specific activities of marker enzymes for amyloplast, cytosol, mitochondria and the vacuole. SDS-PAGE separated amyloplast and starch granule proteins were in-gel digested with trypsin, analyzed by mass spectrometry, and identified by searches against presently available potato protein sequences. Some of these proteins were demonstrated to localize to the amyloplast stroma for the first time. The micro and mini tuber proteomes were very different. However, starch phosphorylase L-1 was particularly abundant in both proteomes. Moreover, disproportionating enzyme 1 and 2, starch phosphorylase L-2 and H, and a-glucan water dikinase were found within the amyloplast, supporting that these specific starch degradative reactions occur within intact potato tuber plastids independently of potential loss of amyloplast envelope integrity during sprouting. The majority of the identified proteins had a predicted plastid transit peptide supporting their presence in the amyloplast. Remarkably, catechol oxidase and transketolase showed twin-arginine translocation (Tat) motives, and 'putative' deoxynucleoside kinase a Sec secretory motif targeting for thylakoid or thylakoid-like structures not expected in amyloplast.