"Lysine is the Lord", thought some scientists in regard to the group interacting with fluorescein isothiocyanate in ATP-binding sites of P-type ATPases but, is it not cysteine?

Isothiocyanates are recognized inhibitors acting on ATP-binding sites of P-type ATPases. Detailed studies with modification of proteins in molecules of purified ATPases by fluorescein isothiocyanate (FITC) and consequent tryptic hydrolysis followed by isolation and sequencing of the respective peptide fragments revealed FITC bound to a lysine residue. This residue was then indicated to be essential for the interaction of ATP with the P-type ATPases. Nevertheless, upon an exchange by site directed mutagenesis of lysine, believed to be essential, the expected total inhibition of ATPase activity was missing. In addition, in the case of the plasma membrane Ca2+-ATPase, the residual activity still remained sensitive to FITC. It was attempted to explain the latter finding by hypothetical existence of some other lysine residue essential for the ATPase activity. On the contrary, in our previous studies we have shown that, based on the reactivity of isothiocyanates, the primary target of FITC in P-type ATPases has to be the SH group of a cysteine residue. However, later on, in altered conditions during trypsinolysis and sequencing, FITC may become transferred from its original site of interaction to a lysine residue and this may lead to final identification of the label on a false place. The present study represents all attempt of elucidating the controversy whether it is lysine or cysteine that represents the FITC-sensitive group truly responsible for the recognition by the active site of P-type ATPases of ATP and its binding.