Isolation and characterization of precursors in bacteriophage T4 baseplate assembly: III. The carboxyl termini of protein P11 are required for assembly activity

Abstract

The assembly activity and electrophoretic mobility of a T4 bacteriophage baseplate protein, P11, have been found to be affected by digestion with the proteases trypsin, subtilisin and carboxypeptidase Y. Analysis of the trypsin limit-digestion product of P11 by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and size analysis by high performance liquid chromatography indicate that there is a decrease of approximately 5000 in the molecular weight of the P11 molecule or a loss of 2500 in M r from each of the gp11 subunits of the dimer. During protease treatment P11 demonstrates a time-dependent loss in the ability to interact with the baseplate protein P10 to form the P( 10 11 ) complex, the first assembly intermediate of the T4 baseplate 1 6 th arm. Similar treatments of the P( 10 11 ) complex indicate that P11 in the complex is not affected by these proteases. Concomitant with the loss of assembly activity is a change in the electrophoretic mobility of P11 on non-denaturing polyacrylamide gels from a single band to a series of more mobile bands suggesting sequential loss of positive charge. P11 assembly activity is completely lost after removal of the first positive charge. These results suggest that the carboxyl termini of the two gp11 subunits of the P11 molecule are involved in the interaction of P11 with P10 to form the P( 10 11 ) complex. Analysis of the portion of gp11 removed by carboxypeptidase Y demonstrates that there are up to 13 aliphatic and aromatic carboxyl-terminal amino acids.