The thiol‐based functionalization of multilayer MoS₂ is performed to customize its electrical and optical performance. Two types of thiol‐terminated organic molecules are utilized: 4‐amino thiophenol (4ATP) for electron‐donating and 4‐nitro thiophenol (4NTP) for electron‐withdrawing. The thiol groups in these molecules chemically bond with MoS₂ at the sulfur vacancies through the simple solution process. 4ATP‐functionalized MoS₂ (4ATP–MoS₂) transistors exhibit a huge enhancement in the current and the carrier concentration, whereas 4NTP‐functionalized MoS₂ (4NTP–MoS₂) transistors show the reduction in the film conductance. Moreover, functionalization with 4ATP and 4NTP complementarily modulate the photoresponsive characteristics of MoS₂ transistors. Under 405 nm laser illumination, 4ATP–MoS₂ phototransistors exhibit the enhanced photoresponsivity of 669.10 A W⁻¹ attributed to the large dark currents. In contrast, 4NTP–MoS₂ phototransistors can achieve faster photoswitching performances than 4ATP–MoS₂. Therefore, the thiol‐based functionalization can effectively customize the electrical and optical characteristics of multilayer MoS₂.