The yeast his4 multifunctional protein. Immunochemistry of the wild type protein and altered forms

A procedure for rapid purification of the yeast his4 protein has been developed. A combined biochemical and immunological study of the his4 proteins from wild type and a number of mutant strains indicates that the his4 gene product is a trifunctional protein with three independent functional domains. Active fragments of the his4 protein possessing one or several activities can result from proteolysis or from genetic alteration. Such fragments can be purified to homogeneity by electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate. Immunological experiments with antibody to the wild type protein and with antibody to genetically or proteolytically altered forms indicate that the sole product of the wild type his4 region is a 95,000-dalton protein. Furthermore, such data indicates that each mutant protein has a characteristic stability that correlates with the type of mutation and with the location of the mutation within the his4 locus. By varying the ionic strength of the extraction buffer, it is possible to find conditions under which such mutant proteins are quite stable.