Spike S1 domain interactome in non-pulmonary systems: A role beyond the receptor recognition

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes Coronavirus Disease 2019 (COVID-19), which, since 2019 in China, has rapidly become a worldwide pandemic. The aggressiveness and global spread were enhanced by the many SARS-CoV-2 variants that have been isolated up to now. These mutations affect mostly the viral glycoprotein Spike (S), the capsid protein mainly involved in the early stages of viral entry processes, through the recognition of specific receptors on the host cell surface. In particular, the subunit S1 of the Spike glycoprotein contains the Receptor Binding Domain (RBD) and it is responsible for the interaction with the angiotensin-converting enzyme 2 (ACE2). Although ACE2 is the primary Spike host receptor currently studied, it has been demonstrated that SARS-CoV-2 is also able to infect cells expressing low levels of ACE2, indicating that the virus may have alternative receptors on the host cells. The identification of the alternative receptors can better elucidate the pathogenicity and the tropism of SARS-CoV-2. Therefore, we investigated the Spike S1 interactomes, starting from host membrane proteins of non-pulmonary cell lines, such as human kidney (HK-2), normal colon (NCM460D), and colorectal adenocarcinoma (Caco-2). We employed an affinity purification-mass spectrometry (AP-MS) to pull down, from the membrane protein extracts of all cell lines, the protein partners of the recombinant form of the Spike S1 domain. The purified interactors were identified by a shotgun proteomics approach. The lists of S1 potential interacting proteins were then clusterized according to cellular localization, biological processes, and pathways, highlighting new possible S1 intracellular functions, crucial not only for the entrance mechanisms but also for viral replication and propagation processes.

Proteomic signature profiling in the cortex of dairy cattle unravels the physiology of brain aging

“…The cortex region was taken post-mortem and analyzed by a comparative label-free proteomics workflow, as reported in Figure 1 . Proteins extracted from each cerebral cortex sample were subjected to a shotgun protocol relying on trypsin hydrolysis by using S-Trap cartridges ( Iacobucci et al, 2022 ). Peptide mixtures were then analyzed by LC–MS/MS, and the raw data was processed by MaxQuant for protein identification and quantification.…”

Preliminary evaluation of the proteomic profiling in the hippocampus of aged grazing cattle

“…Peptide mixtures were dissolved in 100 μL of formic acid 0.2% and 2 μL were analyzed on an LTQ Orbitrap XL (Thermo Fisher Scientific) coupled to the nanoACQUITY UPLC system (Waters; Bertini et al, 2021 ). The software MaxQuant (v. 1.5.2.8) was employed for protein identification and quantification through raw files using the following parameters: UniProt Database; Taxonomy: Bos taurus (Bovine); enzyme: trypsin; 1 missed cleavages allowed; fixed changes: carbamidomethylation (C); variable changes: oxidation (M); Gln-pyro-Glu (N-Term Q); minimum number of peptides for identification: four with at least two unique; minimum number of peptides for quantification four; FDR: 0.01; peptide tolerance: 20 ppm ( Iacobucci et al, 2022 ). Data are available via ProteomeXchange with identifier PXD044404.…”

From the Discovery of Targets to Delivery Systems: How to Decipher and Improve the Metallodrugs’ Actions at a Molecular Level

“…This strategy is also referred to as drug pulldown [ 46 , 47 ]. Pulldown strategies and, more generally, affinity purification-mass spectrometry approaches (AP-MS), are proteomic techniques that allow us to isolate multiprotein complexes starting from a known molecule to hypothesize the intracellular processes in which it is involved [ 48 , 49 , 50 ].…”