Differential proteolysis of the subunits of pyrophosphate-dependent 6-phosphofructo-1-phosphotransferase.
Antibodies against the alpha (Mr 67,000) and beta (Mr 60,000) subunits of wheat seedling Fru-2,6-P2-stimulated pyrophosphate-dependent 6-phosphofructo-1-phosphotransferase (PFP) were used to probe the subunit structures of several partially purified plant PFPs after tryptic digestion. Antisera to the alpha and beta subunits of wheat seedling PFP cross-reacted with the corresponding alpha and beta subunits of PFP preparations from wheat germ, potato tubers, and lettuce leaves. With the mung bean PFP, both antisera reacted with a protein band of Mr 60,000. A protein band corresponding to the Mr 67,000 alpha subunit was not detected in the mung bean PFP preparation. Tryptic digestion of wheat seedling and potato tuber PFPs resulted in the preferential cleavage of the alpha subunit. The trypsinized PFP retained most of its Fru-2,6-P2-stimulated activity but not its basal activity. The proteolyzed enzyme also exhibited a 2-fold increase in Ka for Fru-2,6-P2. Studies with the mung bean enzyme revealed that the anti-alpha immunoreactive component was more sensitive to trypsinization than the anti-beta immunoreactive component of the Mr 60,000 protein band. Thus, the Mr 60,000 protein band of the mung bean PFP appears to be heterogeneous and contains both alpha and beta-like proteins. The above observations indicate that the alpha and beta subunits of PFP are two distinct polypeptides and that alpha acts as a regulatory protein in regulating both the catalytic activity and the Fru-2,6-P2-binding affinity of the beta subunit.
Cheng
H F
HF
Department of Biochemistry, University of Illinois College of Medicine, Chicago 60612.
Tao
M
M
D000906:Antibodies; D000936:Antigen-Antibody Complex; D004591:Electrophoresis, Polyacrylamide Gel; D046911:Macromolecular Substances; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D010770:Phosphotransferases / Q000378:metabolism*; D010944:Plants / Q000201:enzymology*; D011198:Solanum tuberosum / Q000201:enzymology; D014908:Triticum / Q000201:enzymology; D014357:Trypsin
D000906:Antibodies; D000936:Antigen-Antibody Complex; D046911:Macromolecular Substances; D010446:Peptide Fragments; D010770:Phosphotransferases; C022502:pyrophosphate-fructose 6-phosphate 1-phosphotransferase; D014357:Trypsin