Non-enzymatic phosphorylation of bovine serum albumin by Cr(V) complexes: role in Cr(VI)-induced phosphorylation and toxicity.
Evidence for the non-enzymatic phosphorylation of bovine serum albumin (BSA) by sodium bis(2-ethyl-2-hydroxybutyrato)oxochromate(V), Na[CrVO(ehba)2], 1, sodium bis(2-hydroxy-2-methylbutyrato)oxochromate(V), Na[CrVO(hmba)2], 2 and potassium dichromate, K2Cr2O7, 3 in the presence of labeled adenosine-5'-triphosphate (ATP) under conditions of physiological pH is presented. Aggregation and extent of phosphorylation of BSA mediated by 1, 2 or 3 seems to increase with the concentration and time of incubation of the reaction mixture containing all the reactants. The [gamma-32P] label in ATP is incorporated into aggregates of BSA in the in vitro reaction of the protein with ATP in the presence of 1, 2 or 3. Phosphorylation of BSA by ATP in the absence of 1, 2 or 3 is negligible. Addition of EDTA reverses aggregation of protein and liberates partially the incorporated phosphate label. The stoichiometry of phosphorylation is found to be the highest and is equal to 12.25 mol PO4(3-)/mol BSA in the presence of 500 microM of 1, which decreases to 10.56 mol PO4(3-)/mol BSA after EDTA treatment. Resistance to the removal of phosphate label by EDTA increases with increase in time of incubation. Dialysis of phosphorylated BSA reverses the incorporated [gamma-(32)P] label only partially, indicating the formation of covalent links of phosphate groups to BSA. Evidence for the site of phosphorylation in the reaction mediated by 1, 2 or 3 being hydroxyl side groups of tyrosine and serine/threonine residues has been gained. Based on the results, a possibility that 1, 2 and 3 mimic the function of tyrosine and serine/threonine kinases has been invoked.
Vasant
Chellappa
C
Biochemistry Laboratory, Central Leather Research Institute, Chennai 600020, India.
Sankaramanivel
Sundararaj
S
Jana
Mahadevan
M
Rajaram
Rama
R
Ramasami
Thirumalachari
T
D000255:Adenosine Triphosphate / Q000378:metabolism; D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002417:Cattle; D002857:Chromium / Q000378:metabolism* / Q000633:toxicity*; D017608:Chromium Compounds / Q000378:metabolism / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008956:Models, Chemical; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D017382:Reactive Oxygen Species / Q000378:metabolism; D012588:Scintillation Counting; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism
D000596:Amino Acids; D017608:Chromium Compounds; D010761:Phosphorus Radioisotopes; D017382:Reactive Oxygen Species; D002857:Chromium; D012710:Serum Albumin, Bovine; D004533:Egtazic Acid; D000255:Adenosine Triphosphate; D019061:src-Family Kinases