RNA-dependent RNA polymerase: Structure, mechanism, and drug discovery for COVID-19

Abstract

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has rapidly become a global pandemic. Although great efforts have been made to develop effective therapeutic interventions, only the nucleotide analog remdesivir was approved for emergency use against COVID-19. Remdesivir targets the RNA-dependent RNA polymerase (RdRp), an essential enzyme for viral RNA replication and a promising drug target for COVID-19. Recently, several structures of RdRp in complex with substrate RNA and remdesivir were reported, providing insights into the mechanisms of RNA recognition by RdRp. These structures also reveal the mechanism of RdRp inhibition by nucleotide inhibitors and offer a molecular template for the development of RdRp-targeting drugs. This review discusses the recognition mechanism of RNA and nucleotide inhibitor by RdRp, and its implication in drug discovery.

Keywords: COVID-19; RNA recognition; RNA-dependent RNA polymerase; Remdesivir; SARS-CoV-2; Structure.

Fig. 1

The genome of SARS-CoV-2. (A) The organization of SARS-CoV-2 genome. (B) The polyproteins (pp1a and pp1ab) are cleaved into 16 non-structural proteins (nsp1-16).

Fig. 2

Structure of the SARS-CoV-2 apo RdRp complex. (A) The schematic diagram for the domain organization of the RdRp complex, containing nsp12, nsp7, and two copies of nsp8 (nsp8-1 and nsp8-2). The polymerase motifs A to G in the catalytic site are highlighted. The β-hairpin is indicated. (B–C) Two views of the cryo-EM map (B) and structure (C) of SARS-CoV-2 apo RdRp complex (PDB code: 7BV1). (D) The conserved zinc-binding motifs. The zinc-binding residues are shown as sticks. The subdomains and components of the RdRp complex are colored as follows: β-hairpin, chocolate; NiRAN, gold; Interface, tomato; Fingers, salmon; Palm, green; Thumb, light blue; nsp7, magenta; nsp8-1, dark cyan; nsp8-2, cyan. (E) The active site of the RdRp complex. The conserved seven motifs A-G are highlighted as indicated colors. The conserved residues, K545 and R555 in motif F, D618 and D623 in motif A, as well as D760 and D761 in motif C, are shown as sticks. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 3

The structural basis for RNA recognition by SARS-CoV-2 RdRp. (A) Two surface views of the RdRp with the electrostatic potential (Red, negative; blue, positive). The primer-template entry, product exit, and nucleotide (NTP) entry routes are highlighted. (B) Superposition of structures of apo (PDB codes: 7BV1) and RNA bound RdRp complex (PDB codes: 7BV2). The subdomains of the RNA bound RdRp complex are colored as indicated, while that of the apo RdRp complex are shown in gray. (C) Structural rearrangements of RdRp active site after RNA binding. The loops in motif B, motif G, and the thumb subdomain in RNA bound RdRp structure are colored in cyan, salmon, and light blue, respectively, while corresponding loops in apo RdRp structure are indicated in pale cyan, light salmon, and gray. The directions of red arrows indicate the shift of the structural fragments from apo RdRp toward that from RNA bound RdRp complex. (D–E) The “sliding poles”-like conformation of the N-terminal extension of nsp8 stabilizes the long exiting helical RNA (PDB codes: 6YYT). The subdomains of the RdRp complex, and the template and product strands are colored as indicated. The solid circled numbers 1 to 11 indicate the positively charged residues at the N-terminal of nsp8, including K36, K37, K39, K40, N43, K46, R51, R57, K58, and Q65, respectively (E). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4

Inhibition mechanism of RdRp by remdesivir. (A) Chemical structure of remdesivir and its cellular metabolic pathway. (B) The binding mode of remdesivir in the active catalytic site of SARS-CoV-2 RdRp (PDB code: 7BV2). The covalently bound remdesivir in the monophosphate form (RDV-MP, colored in magenta), two magnesium ions (green), and pyrophosphate (orange) are shown. Residues interacted with RDV-MP in motif A (green), motif B (cyan), motif C (chocolate), and motif F (light blue) are shown as sticks. Polar interactions formed between RDV-MP and primer strand (yellow orange), as well as template strand (orange) are highlighted as red dashed lines. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)