The effects of Ni promoter on Mo-S/CNT catalyst towards hydrodesulfurization (HDS) and hydrogen evolution reactions (HER) were studied. Ni promoter was introduced into Mo-S/CNT catalyst by one-step or mechanical mixing method, and Ni species presented in the form of Ni-Mo-S or Ni3S4, respectively. Catalytic assessments indicated that Ni-Mo-S structure had obviously better HDS and HER activities than MoS2 at the same time, while Ni3S4 could not improve the activity of MoS2 towards HDS or HER. The density functional theory (DFT) was also used to simulate and calculate the activation energies needed for HDS and HER on MoS2 structure and Ni-Mo-S structure, respectively. It was found that the required activation energies for C-S bond breaking and H-2 generation on Ni-Mo-S structure were both significantly lower than that of MoS2 structure. Therefore, it was proved theoretically that compared with MoS2 structure, Ni-Mo-S structure had better catalytic activities for both HDS and HER.