High Dihydrofolate Reductase Levels in Diplococcus pneumoniae After Mutation in the Structural Gene: Biochemical and Immunological Evidence for Increased Synthesis

Abstract

A unique class of missense mutations in the dihydrofolate reductase structural gene of Diplococcus pneumoniae determines marked increases in the corresponding enzyme activity. Immunological evidence, based on precipitating and neutralizing antibody reactions, is presented in support of previously derived findings which attribute this increase in activity to an increase in the amount of enzyme protein.Biochemical evidence was obtained which shows that the increased amounts of dihydrofolate reductase protein in the mutants not owing to a decrease in degradation, reducing turnover, or to an increase in stability, and, therefore, probably related to an increase in the rate of synthesis of new protein. The manner in which a missense alteration of a structural gene might quantitatively alter the expression of that gene is discussed.Immunoassay. Antibody against dihydrofolate reductase was prepared in a female albino rabbit by two injections, 10 days apart, of 0.5 mg of purified dihydrofolate reductase protein in I ml of 0.02 M 318 on July 16, 2020 by guest http://jb.asm.org/ Downloaded from VOL. 106, 1971 DIHYDROFOLATE REDUCTASE LEVELS IN D. PNEUMONIAE 320 SIROTNAK on July 16, 2020 by guest