Distribution of separations between groups in an engineered calmodulin

An engineered calmodulin (VU-9-CaM) has been prepared in which a tryptophan group is present at position 99 and a tyrosine at position 138. The tyrosine was converted to nitrotyrosine. Timedomain dynamic fluorescence measurements were made of energy transfer from the tryptophan donor to the nitrotyrosine acceptor. These were analyzed to yield the parameters characterizing the distribution of separations between the two groups, which are located on Ca²⁺-binding domains III and IV. Their mean separation is in reasonable agreement with the crystallographic value.