Polyoma virus middle T antigen: relationship to cell membranes and apparent lack of ATP-binding activity.
Middle T antigen of polyoma virus is associated principally with the plasma membrane. Comparison of the trypsin sensitivity of middle T in intact cells and "inside out" membrane preparations showed that middle T is oriented towards the inside of the cell. This was confirmed by labeling of middle T in permeabilized cells, but not in intact cells, using [gamma-32P]ATP. Middle T molecules active in the in vitro kinase reaction could be differentiated from the bulk (metabolically labeled) middle T based on resistance to trypsin treatment. The active fraction also behaved differently from the bulk when cell frameworks were prepared with Triton-containing buffers; whereas the bulk middle T was evenly distributed in the soluble and cell framework fractions, the kinase-active forms were largely associated with the framework. Middle T molecules labeled in vivo with 32PO4 were found largely in the framework fraction, like the molecules that show kinase activity in vitro. Experiments with ATP affinity reagents 8-azido-ATP and 2,3-dialdehyde ATP have failed to label the middle T antigen. However, 2,3-dialdehyde ATP could be used to inhibit the kinase reaction. This raises the question of whether middle T antigen possesses intrinsic kinase activity or, rather, associates with a cellular tyrosine kinase.
D000956:Antigens, Viral; D000957:Antigens, Viral, Tumor; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000276:immunology*; D003599:Cytoskeleton / Q000201:enzymology; D008970:Molecular Weight; D019000:Phosphotyrosine; D011120:Polyomavirus / Q000276:immunology*; D011494:Protein Kinases / Q000378:metabolism; D014443:Tyrosine / Q000031:analogs & derivatives / Q000096:biosynthesis
D000956:Antigens, Viral; D000957:Antigens, Viral, Tumor; D019000:Phosphotyrosine; D014443:Tyrosine; D011494:Protein Kinases