Hamm H H, Decker K
Eur J Biochem. 1978 Dec;92(2):449-54. doi: 10.1111/j.1432-1033.1978.tb12766.x.
The process, by which FAD is attached covalently to the 6-hydroxy-D-nicotine oxidase apoprotein in D-nicotine-induced cells of Arthrobacter oxidans was studied in vitro. [3H]Adenine-labelled FAD prepared biosynthetically in Clostridium kluyveri was incorporated into the 6-hydroxy-D-nicotine oxidase molecule during cell-free translation. FAD rather than FMN or riboflavin was thus shown to be the flavin derivative transferred to the polypeptide chain. After short-term protein synthesis on ribosomes from induced A. oxidans cells in the presence of an Escherichia coli MRE 600 supernatant fraction and [adenine-2-3H]FAD, THE PEPTIDYL-TRNA fraction was separated from completed polypeptides. Labelled FAD was found to be covalently attached to the tRNA-bound polypeptides. Cleavage of the tRNA-peptide bond released labelled polypeptides the largest of which migrated as authentic 6-hydroxy-D-nicotine oxidase during dodecylsulfate/polyacrylamide gel electrophoresis. These results strongly suggest that FAD is incorporated into the nascent polypeptide chains of 6-hydroxy-D-nicotine oxidase during ribosomal translation.
