In recent years, numerous researchers have incorporated plant fibers into polymers to alter the thermal and mechanical properties of materials. Grapevines, considered agricultural waste, have led to burdens for farmers and environmental challenges due to their mass production. This study aims to reduce the brittleness of polylactic acid (PLA) by adding polybutylene succinate (PBS) as a toughening agent and employing grapevine fiber (GVF) as a biomass filler. Additionally, the influence of GVF, toughening agents, compatibilizers, and lubrication agents on the tensile strength, heat deflection temperature (HDT), and impact strength of the composites was examined. The findings revealed that the addition of 10% GVF and 5% PBS increased the impact and tensile strengths of PLA from 17.47 J/m and 49.74 MPa to 29.7 J/m and 54.46 MPa, respectively. Moreover, the HDT of the composites exceeded 120 °C when the GVF content was more than 40 wt%. Additionally, the inclusion of a compatibilizer and a lubrication agent enabled the composite containing 30% GVF to achieve tensile and impact strengths of 45.30 MPa and 25.52 J/m, respectively. Consequently, these GVF/PLA green bio-composites not only improve the mechanical and thermal properties of PLA but also promote the reuse of waste grapevines.