Variable Macro X Domain of SARS-CoV-2 Retains the Ability to Bind ADP-ribose

David N. Frick; Rajdeep S. Virdi; Nemanja Vuksanovic; Narayan Dahal; Nicholas R Silvaggi

Many of the 47 variant residues in the 180 amino acid long SARS macro X domain are clustered near its N-terminus in a region that is particularly variable in the three Coronaviridae genera (Fig. 2) . Macro X domains from coronaviruses that cause the common cold (alpha coronaviruses) 5 and the beta coronaviruses like SARS-CoV-1 and Middle East respiratory syndrome coronavirus (MERS-CoV) 8 all bind ADP ribose. However, the same domain in gamma coronaviruses does not bind ADPribose. 5 Expression and Purification of the SARS-CoV-2 Macro X domain-An E. coli expression vector for the macro X domain was built to express and N-terminally His-tagged protein similar to a SARS-CoV-1 protein studied by Egloff et al. 9 Upon induction, a one liter culture of BL21(DE3) cells harboring the vector expresses 50-100 mg of the macro X domain protein that can be purified in one step to apparent homogeneity using immobilized metal affinity chromatography (Fig. 3A) .