key: cord-1051746-gsqd57b6
authors: Benvenuto, Domenico; Demir, Ayse Banu; Giovanetti, Marta; Bianchi, Martina; Angeletti, Silvia; Pascarella, Stefano; Cauda, Roberto; Ciccozzi, Massimo; Cassone, Antonio
title: Evidence for mutations in SARS‐CoV‐2 Italian isolates potentially affecting virus transmission
date: 2020-06-03
journal: J Med Virol
DOI: 10.1002/jmv.26104
sha: 605dfcba18822d26ce42a4a6debe4345cc263838
doc_id: 1051746
cord_uid: gsqd57b6

Italy is the first western country suffering heavy SARS‐CoV‐2 transmission and disease impact after Covid‐19 pandemia started in China. Even though the presence of mutations on spike glycoprotein and nucleocapsid in Italian isolates has been reported, the potential impact of these mutations on viral transmission has not been evaluated. We have compared SARS‐CoV‐2 genome sequences from Italian patients with virus sequences from Chinese patients. We focussed upon three non‐synonimous mutations of genes coding for S(one) and N (two) viral proteins present in Italian isolates and absent in Chinese ones, using various bio‐informatic tools. Amino acid analysis and changes in three‐dimensional protein structure suggests the mutations reduce protein stability and, particularly for S1 mutation, the enhanced torsional ability of the molecule could favour virus binding to cell receptor(s). This theoretical interpretation awaits experimental and clinical confirmation. This article is protected by copyright. All rights reserved.

online server has been used to estimate the stability of potential mutations found using the selective pressure analysis. three-dimensional structures have been analyzed and displayed using PyMOL [12] .

The analysis of the alignment has revealed the presence of two mutations on Spike and Nucleocapsid (N) proteins.

Regarding the Spike glycoprotein, the transition from an Adenosine to Guanine occurring on the 1901st nucleotide has led to non-synonymous mutation from an Aspartate to a Glycine residue in the 614 aminoacidic position (found in all the Italian isolates). This mutation, which was first reported in italian isolates by Zehender et al This article is protected by copyright. All rights reserved.

Article (20) , is characteristic of all the Italian SARS-CoV-2 sequences besides those of the numerous viral isolates in Abruzzo (4). Aspartate is a chiral and polar amino acid which frequently occurs at the N-termini of alpha helices while Glycine is non-polar and the only achiral amino acid. This mutation is located on the SD2 region of the RBDcontaining S1 subunit [13] . Using the crystallographic 3-dimensional structure of the S protein, the implications of this mutation has been analysed using CUPSAT and Dynamut servers. The results of these analysis have shown that the mutation from Aspartate to Glycine reduces the stability of the protein (ΔΔG (kcal/mol) -1.51) favouring the torsion potential. The Δ Vibrational Entropy Energy Between Wild-Type and Mutant Spike protein has been calculated to be: ΔΔSVib ENCoM: 0.065 kcal.mol-1.

K-1. (Figure 1) Regarding the Nucleocapsid protein, the two contiguous mutations (found in 56% of Italian isolates), from AGG to AAA occurring on the 649 -651 nucleotides and GGA to CGA occurring on the 652 -655 nucleotides both lead to non-synonymous mutation from an Arginine to a Lysine and from Glycine to Arginine residue in the 203 and 204 aminoacidic position, respectively. This latter mutation is characteristic of the SARS-CoV-2 sequences isolated in Abruzzo. Arginine is a chiral and polar amino acid which frequently occurs at the N-termini of alpha helices while Lysine is a chiral and polar amino acid (for Glycine, see above). Using the 3-dimensional structure available on Itasser server, the implication of these mutations has been analysed by CUPSAT server.

The results point out both mutations reduce protein stability while favouring torsion (ΔΔG (kcal/mol) -1.92 and -2.94 for arginine to lysine and glycine to arginine, respectively). The homology modelling analysis performed using HHpred server has This article is protected by copyright. All rights reserved. 

Genomic characterization and phylogenetic analysis of SARS-COV-2 in Italy

Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV

APPENDIX While this paper was in advanced preparation for publication