TABLE 3

Functional analysis of Ala replacement mutants

Ime1p derivativeaime2-lacZ expressionb (Miller units)Sporulationb (%)Phosphorylation
No Ime1P1.1<0.1
Ime1Δ68-182P19673+
Ime1Δ68-182-Q340*p4.01.0
Ime1Δ68-182-S352Ap4333+
Ime1Δ68-182-S356Ap8628+
Ime1Δ68-182-S360Ap16746+
Ime1Δ68-182-S352A, S356A, S360Ap6230+
Ime1Δ68-182p (low expression)c14865+
Ime1Δ68-182-ala8p2.0<0.1
  • a Ime1p derivatives are all expressed from the GAL1 promoter. Ime1Δ68-182-ala8p has substitutions Y342A, Y343A, S352A, Y353A, S356A, T358A, Y359A, and S360A.

  • b Strain M1121 (ime1Δ/ime1ΔIME2/ime2-lacZ) carrying the indicated PGAL1-Ime1Δ68-182 plasmids was incubated in sporulation medium for determination of β-galactosidase activity (after 8 hr) and sporulation ability (after 24–72 hr).

  • c Ime1Δ68-182p was expressed at reduced levels from plasmid pKM212, which lacks IME1 3′ noncoding sequences.