In vitro identification of a soluble protein:geranylgeranyl transferase from rat tissues

Area of Science:

• Biochemistry
• Molecular Biology
• Cellular Biology

Background:

• Mammalian G protein gamma subunits undergo in vivo modification via prenylation with a geranylgeranyl group.
• This prenylation occurs at a specific cysteine residue within the carboxyl-terminal sequence.
• Prenylation is a critical post-translational modification influencing protein function and localization.

Purpose of the Study:

• To investigate protein:geranylgeranyl transferase activity in rat organs across different ages.
• To characterize the enzymatic activity, including optimal conditions and substrate specificity.
• To determine the distribution and age-dependent changes of this prenyltransferase activity.

Main Methods:

• Utilized a biotinylated peptide acceptor mimicking the gamma subunit's C-terminus.
• Employed tritiated geranylgeranyl diphosphate as a substrate to assay transferase activity.
• Measured enzyme activity in various rat organs (brain S100, liver, kidney, heart) at different ages, assessing dependence on metal ions, pH, and substrate/protein concentrations.

Main Results:

• Protein:geranylgeranyl transferase activity was detected in all studied rat organs, with the highest specific activity in brain S100.
• Activity was dependent on divalent metal ions (ZnCl2 or MgCl2) and exhibited optimal conditions at pH 7.5.
• Specific activity significantly increased with age in brain, liver, kidney, and heart, and was absent in the membrane fraction.

Conclusions:

• Rat brain S100 possesses both protein:geranylgeranyl transferase and geranylgeranyl synthetase activities.
• The peptide acceptor is specific for geranylgeranyl modification under the tested conditions.
• Geranylgeranyl transferase activity in rat organs, especially brain, shows a notable age-dependent increase.