Specific pupylation as IDEntity reporter(SPIDER)for the identification of protein-biomolecule interactions

Protein-biomolecule interactions play pivotal roles in almost all biological processes.For a biomolecule of interest,the identification of the interacting protein(s)is essential.For this need,although many assays are available,highly robust and reliable methods are always desired.By combining a substrate-based proximity labeling activity from the pupylation pathway of Mycobacterium tuberculosis and the streptavidin(SA)-biotin system,we developed the Specific Pupylation as IDEntity Reporter(SPIDER)method for identifying protein-biomolecule interactions.Using SPIDER,we validated the interactions between the known binding proteins of protein,DNA,RNA,and small molecule.We successfully applied SPIDER to construct the global protein interactome for m^(6)A and m RNA,identified a variety of uncharacterized m^(6)A binding proteins,and validated SRSF7 as a potential m^(6)A reader.We globally identified the binding proteins for lenalidomide and Cob B.Moreover,we identified SARS-CoV-2-specific receptors on the cell membrane.Overall,SPIDER is powerful and highly accessible for the study of proteinbiomolecule interactions.

Systematic evaluation of IgG responses to SARS-CoV-2 spike protein-derived peptides for monitoring COVID-19 patients

Serological tests play an essential role in monitoring and combating the COVID-19 pandemic.Recombinant spike protein(S protein),especially the S1 protein,is one of the major reagents used for serological tests.However,the high cost of S protein production and possible cross-reactivity with other human coronaviruses pose unavoidable challenges.By taking advantage of a peptide microarray with full spike protein coverage,we analyzed 2,434 sera from 858 COVID-19 patients,63 asymptomatic patients and 610 controls collected from multiple clinical centers.Based on the results,we identified several S protein-derived 12-mer peptides that have high diagnostic performance.In particular,for monitoring the IgG response,one peptide(aa 1148-1159 or S2-78)exhibited a sensitivity(95.5%,95%CI 93.7-96.9%)and specificity(96.7%,95%CI 94.8-98.0%)comparable to those of the S1 protein for the detection of both symptomatic and asymptomatic COVID-19 cases.Furthermore,the diagnostic performance of the S2-78(aa 1148-1159)IgG was successfully validated by ELISA in an independent sample cohort.A panel of four peptides,S1-93(aa 553-564),S1-97(aa 577-588),S1-101(aa 601-612)and S1-105(aa 625-636),that likely will avoid potential cross-reactivity with sera from patients infected by other coronaviruses was constructed.The peptides identified in this study may be applied independently or in combination with the S1 protein for accurate,affordable,and accessible COVID-19 diagnosis.