In this study, we have performed MD simulations to elucidate the molecular mechanisms for the "template-dependent inhibition" exerted by Remdesivir. We first examined if the NTP incorporation at the active site is directly impaired when Remdesivir at +1 site (Fig. and). The cryo-EM structure of the SARS-CoV-2 nsp12-nsp7-nsp8 complex (PDBID:7BZF ) was adopted as our structural basis and Remdesivir was model at +1 site of the template strand with UTP binding at a closed active site (see SI Section 1.2 for details of model construction). Five replicas of 50 ns production simulations have been performed to investigate the incorporation capability of UTP opposite to Remdesivir (Fig. ). For comparison, the same amount of MD simulations with UTP opposite to adenosine ("wildtype-RNA" system, Fig. ) has also been conducted (see SI Section 1.1 for details). The distance between the Pα atom of UTP and the O3' atom of the 3'-terminal of nascent strand (Pα-O3' distance, Fig. ) below 4 Å has been suggested to be necessary for efficient catalysis and formation of phosphodiester bond . Consistently, such distance is <4 Å in all the MD conformations with a UTP:A pair at the active site (Fig. ). Interestingly, when Remdesivir is present at the template strand complementary to UTP, all the MD simulations still retain the Pα-O3' distance below 4 Å (Fig. ), which satisfies the prerequisite for the efficient incorporation. The stability of UTP at the active site is also important for ensuring a catalytically active configuration. Accordingly, the base pairing and base stacking stability were examined (Fig. and). Our results demonstrated that UTP can always form stable base pairing with Remdesivir embedded at +1 site of the template strand (hydrogen bonding probability = 95.6%), resembling that for the UTP:A base pair (hydrogen bonding probability = 93.6%) (Fig. ). Moreover, UTP can stack well with the 3'-terminal of the nascent strand in both systems, with the base-to-base dihedral angles equal to 19.9° and 20.3° for adenosine and Remdesivir, respectively (Fig. ). Altogether, our calculations have suggested that UTP could well maintain a catalytically active configuration opposite to Remdesivir at +1 site of the template strand.