[9] Determination of GTP, GDP, and GMP in cell and tissue extracts

This chapter describes a simple, sensitive, and reproducible method for determining the concentrations of guanosine triphosphate (GTP), guanosine diphosphate (GDP), and guanosine monophosphate (GMP) in extracts of biological materials, without prior chromatographic separations. This procedure is based upon the selective enzymic degradation of specific nucleotide triphosphates (NTPs) within cell extracts, and the fact that crude firefly-luciferase preparations contain transphosphorylase enzymes and adenosine diphosphate (ADP), which together result in the quantitative production of adenosine triphosphate (ATP) in response to the addition of a wide variety of ribose and deoxyribose nucleotide triphosphates. A complete description of the experimental procedures and equipment necessary for the determination of GTP, GDP, and GMP, the pertinent analytical principles, assay limitations, and possible sources of interference are presented in the chapter.