The critical current density (Jc) of Y0.5Gd0.5Ba2Cu3O7-x (YGdBCO)-coated conductors (CCs) could be improved via producing the periodic micro-hole arrays on the surface of CCs. In this work, the influence of micro-hole arrays on the transport AC losses of YGdBCO CCs was studied. The micro-hole arrays were produced in YGdBCO CCs via laser drilling, which were distributed with different spacings (l) between two neighboring micro-holes along longitudinal and transverse directions. With l ≥ 0.7 × 0.7 mm, the critical current (Ic) values of drilled samples were nearly identical with that of virgin one. However, a considerable nonlinear decrease of Ic value was displayed in the drilled samples with l < 0.7 × 0.7 mm. In addition, compared to the virgin one, the AC losses of samples drilled with various transverse and longitudinal spacings were increased more than one order of magnitude under low frequency (31 Hz ~ 177 Hz). There was almost no effect on the AC losses of micro-holes spacing for drilled CCs. A 3D H-formulation simulation model was established to analyze the experimental results. The investigations revealed the magnetic flux densities around micro-holes were altered, which could potentially lead to an increase of AC loss.