FRACTIONATION AND CHARACTERIZATION OF PROTEIN C-TERMINAL PRENYLCYSTEINE METHYLESTERASE ACTIVITIES FROM RABBIT BRAIN

Reversible carboxyl methylation of the C-terminal geranylgeranylcystei ne of G25K may regulate its activity and cellular localization. Brain homogenates were examined for enzyme activities which hydrolyze the me thyl ester of [H-3]methyl-G25K to produce [H-3]methanol. Methylesteras e activity was detected in both soluble and membrane fractions. The so luble activity was fractionated into at least two distinct activities. One soluble activity appears to be due to the lysosomal protease, cat hepsin B, based on sensitivity to certain protease inhibitors, acidic pH optimum, size, and ability to cleave the peptide substrate N alpha- CBZ-Arg-Arg-7 amido-4-methylcoumarin. A second soluble activity, assoc iated with a protein of approximately 25 kDa, exhibits a neutral pH op timum, insensitivity to protease inhibitors, and inhibition by the est erase inhibitor, ebelactone B. The membrane fraction contains larger a mounts of a similar methylesterase that may represent the physiologica lly relevant form of the enzyme. (C) 1995 Academic Press, Inc.