KLP-6 is a kinesin superfamily protein resistant to ADP inhibition

Abstract Product inhibition is a type of enzyme inhibition in which the reaction product suppresses further enzyme activity. Kinesins are microtubule-dependent ATPases that move along microtubules by hydrolyzing ATP into ADP and inorganic phosphate (Pi). Like other ATPases, kinesins are inhibited by their hydrolysis product, ADP. We show here that Caenorhabditis elegans kinesin-3, KLP-6, is a unique kinesin that is resistant to ADP inhibition. Amino acid sequence comparisons between KLP-6 and KIF1A, another kinesin-3, revealed that KLP-6 possesses a unique sequence in an α2a helix. Substituting this region with the corresponding sequence from KIF1A abolished KLP-6’s markedly higher binding selectivity for ATP over ADP, confirming that this domain is crucial for its insensitivity to ADP inhibition. Molecular dynamics simulations uncovered that the α2a helix of KLP-6 adopts a less stable helical conformation in the ADP-bound state than in the ATP-bound state, which explains KLP-6’s strong selectivity for ATP. Our results provide valuable insights into kinesin’s nucleotide selectivity and underlying molecular mechanism.