Display omitted •UiO-66-NH2 with different kinds of defects were synthesized.•UiO-66-NH2-LV with ligand vacancy exhibits the best photocatalytic activity.•DFT calculations reveal UiO-66-NH2-LV has the lowest sum of Eabs and ELMCT in the photocatalytic process. Defects in MOFs can benefit light absorption and charge transfer for photocatalytic application, nevertheless, studies on interplay between structural defects and photocatalytic properties of MOFs are in infancy. Herein, a series of UiO-66-NH2 with different kinds of defects were created for regulating CO2 photocatalytic reduction. Theoretical calculations in combination with experimental data verified that the sample with ligand-vacant (UiO-66-NH2-LV) defect performed better than non-defect, missing-cluster and monocarboxylate compensated ones in the photocatalytic CO2 reduction reactions. UiO-66-NH2-LV shows superior photocatalytic activity with the highest CO yield of 30.5 μmol g−1h−1, which is 9.2 times higher than that of the sample with missing-cluster (UiO-66-NH2-MC), as well as the highest quantum yield (QY) of 0.90%. DFT calculations further demonstrate the correlation between discriminative photocatalytic activities in defect structures and tunable electronic properties characterized by absorption energy, Eabs, and charge transfer energy, ELMCT, in the photocatalytic process. The ligand-vacant defect with the lowest sum of Eabs and ELMCT will lower photocatalytic reaction energy barrier in the rate-limiting step among the elementary reaction step during CO2 photoreduction. The insights gained from this study will guide the MOFs defect-engineering for enhancing CO2 photocatalytic capacity.