35S-labelled thiophosphorylated derivative of inositol trisphosphate

We have prepared the [35S]thiophosphate labelled inositol trisphosphate and [32P]phosphate labelled inositol trisphosphate (IP3) from human erythrocytes. These compounds were used as substrates for the inositol trisphosphate 5-phosphomonesterase assay in human erythrocyte membranes. During 60 min incubation with the enzyme, the 35S-labelled IP3 was not hydrolyzed, meanwhile the 32P-labelled IP3 was broken down to 19.8 ± 2.4%. This suggests that due to the presence of thiophosphate in the molecule the 35S-labelled IP3 cannot serve as substrate for inositol trisphosphate 5-phosphomonoesterase. The [35S]thiophosphorylated derivative thus represents a nonhydrolyzable analogue of IP3 and could be used in the study of its second messenger role. (author)