The Los Burros gold deposit is located within the Franciscan complex of the Nacimiento block, California. Although this deposit was mined during the late 19th and early 20th centuries, its origin and its relationship with the regional metamorphism and deformation are unclear. In order to better understand the mode of formation of the deposit, and its possible link with a post-subduction thermal anomaly, we performed a detailed geological and structural mapping within the Los Burros district, coupled with a (U-Th)/He thermochronological study on apatite and zircon. Fluid-inclusion and stable isotope (O and C) analyses of the ore deposit provide constraints on the temperature and potential source of the fluid(s) responsible for the Los Burros gold formation. Our results demonstrate that the Los Burros Au deposit formed from a metamorphic aqueous CO2-CH4 fluid at temperatures of ~300 °C. Results from Raman Spectroscopy of Carbonaceous Materials confirm the presence of a thermal anomaly in the range 260–320 °C near the Los Burros Mining District as previously inferred by vitrinite reflectance techniques. Our structural and preliminary thermochronological analyses show that the Los Burros Au deposit consists of a series of meter-wide, quartz-calcite-sulfide “en-echelon” veins, formed during a late transpressive event related to San Andreas fault activity and the subduction of a slab-window during the past 30 Ma. Our new data suggest that the thermal anomaly recorded in the vicinity of the Los Burros Mining District is not associated with a magmatic-derived hydrothermal system such as described within the epithermal McLaughlin deposit in Northern California, but rather with a late transpressive deformation event that uplifted deeply buried metamorphic rocks, possibly connected to a local metamorphic overprint.