Maturation of the tail spike endorhamnosidase of Salmonella phage P22.

As part of a genetic analysis of the in vivo folding and subunit assembly of the P22 tail spike endorhamnosidase, we have studied the maturation of the newly synthesized 76,000-dalton polypeptide chains into thermostable tail spike oligomers. Four of 15 temperature-sensitive mutations in the structural gene for this protein result in electrophoretically distinct tail spikes. Cells mixedly infected with wild type and an electrophoretic variant produce two hybrid species, with mobilities intermediate between the parental species, indicating that the native tail spike is a trimer. Mature trimers are resistant to denaturation by sodium dodecyl sulfate (SDS): at room temperature the trimer migrates in an SDS gel as if it were not binding significant amounts of SDS, whereas the heat-denatured chain migrates as expected of an SDS-polypeptide complex. The mature trimer is also resistant to trypsin digestion. Lysates of infected cells contain SDS and trypsin-sensitive forms of the newly synthesized tail spike polypeptide chains. These are probably incompletely or incorrectly folded chains. SDS and trypsin resistance were used to measure the efficiency of in vivo folding and subunit assembly of the mature trimer from its polypeptide chains. This decreased from 90% at 27 degrees C to only 15% at 42 degrees C. These results are consistent with the existence or a labile intermediate or step in the folding or subunit assembly of the thermostable tail spike protein. We discuss the possibility that the achievement of certain structural features of mature proteins may entail difficulties in their folding pathways.