ABSTRACT
Goldenberg, DP and King, J. (1981) J. Mol Biol., 145(4), 633-651
Temperature-sensitive mutants blocked in the folding or subunit of the bacteriophage P22 tailspike protein. II. Active mutant proteins matured at 30° C
Little is known of the molecular mechanisms by which temperature-sensitive mutatins interfere with the formation of biologically active proteins. We have studied the effects of such mutations at 13 different sites on the properties of the mutifunctional tail spike protein of bacteriophage P22, a thermostable structural protein composed of 76,000 Mr chains.
Using multiple mutant strains blocked in capsid assembly, we have examined the free mutant tail spikes that accumulate in active form at permissive temperature. When assayed for the ability to bind to phage heads at the restrictive temperature, the mutant proteins were as active as the wild type. Similarly, when assayed for the ability to adsorb to bacteria at restrictive temperature, the mutant proteins were as active as the wild type. Thus the temperature-sensitive phenotypes of the mutants are not due to the thermolability of these functions in the mature mutant protein.
The wild-type protein is heat-resistant, requiring incubation at 90°C, to give a halt-time of inactivation of ten minutes. The 13 ts mutant proteins, once matured at 30°C and 39°C is the same, but the yield of active tail spikes at 39°C is only 25% of the yield at 30°C.
The results show that the amino acid substitutions in the mutant proteins, though lethal for the formation of the virus at 39°C, do not affect the thermostability of the mature tail spike protein formed at 30°C. They may act by destabilizing thermolabile intermediates in the folding or subunit assembly of the tail spike protein.
