Reversible compartmentalization of de novo purine biosynthetic complexes in living cells.
Purines are synthesized de novo in 10 chemical steps that are catalyzed by six enzymes in eukaryotes. Studies in vitro have provided little evidence of anticipated protein-protein interactions that would enable substrate channeling and regulation of the metabolic flux. We applied fluorescence microscopy to HeLa cells and discovered that all six enzymes colocalize to form clusters in the cellular cytoplasm. The association and dissociation of these enzyme clusters can be regulated dynamically, by either changing the purine levels of or adding exogenous agents to the culture media. Collectively, the data provide strong evidence for the formation of a multi-enzyme complex, the "purinosome," to carry out de novo purine biosynthesis in cells.
An
Songon
S
Department of Chemistry, Pennsylvania State University, University Park, PA 16802, USA. sua13@psu.edu
Kumar
Ravindra
R
Sheets
Erin D
ED
Benkovic
Stephen J
SJ
D001377:Azaserine / Q000494:pharmacology; D001665:Binding Sites; D019731:Carbon-Nitrogen Ligases / Q000235:genetics / Q000378:metabolism*; D019733:Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor / Q000235:genetics / Q000378:metabolism*; D002451:Cell Compartmentation; D002460:Cell Line; D045744:Cell Line, Tumor; D003470:Culture Media; D003593:Cytoplasm / Q000201:enzymology*; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D019271:Hypoxanthine / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism*; D051026:Phosphoribosylglycinamide Formyltransferase / Q000235:genetics / Q000378:metabolism*; D011687:Purines / Q000096:biosynthesis*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
D003470:Culture Media; D009097:Multienzyme Complexes; D011687:Purines; D011993:Recombinant Fusion Proteins; D019271:Hypoxanthine; D001377:Azaserine; D051026:Phosphoribosylglycinamide Formyltransferase; D019731:Carbon-Nitrogen Ligases; C022380:phosphoribosylaminoimidazole synthase; C022387:phosphoribosylamine-glycine ligase; D019733:Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor; C022385:phosphoribosylformylglycinamidine synthetase