Kinesin-3 KIF1A is responsible for the anterograde transport of synapse vesicle (SV) precursors in axons. The CC1-FHA tandem of KIF1A has been revealed as a stable dimer that can trigger motor activity, but the mechanism underlying the regulation of the CC1-FHA dimer is unclear. Based on the CC1-FHA dimer structure, we found a potential phosphorylation motif "⁴⁸⁷SPKK⁴⁹⁰" located at the dimer interface. We demonstrated that the phosphorylation-mimetic mutation of Ser487 leads to the dissociation of the CC1-FHA dimer. Moreover, the Ser487-mutation could regulate the motor activity of KIF1A and the KIF1A-mediated axonal transport activity of SVs in C. elegans. Thus, the highly conserved "⁴⁸⁷SPKK⁴⁹⁰" motif may be a key site in the CC1-FHA tandem for regulating CC1-FHA dimerization and the subsequent activity of KIF1A.