# Functionnal annotation - Example analysis of S-protein polymorphism

# Live Resources

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# What's the point?

In the previous portion of this study, we found variations in SARS-2 Cov genome. To evaluate the impact of variations on the virus, we need to perform functional annotation of variants. A lot of literature is available on a wide variety of species of coronaviruses. To help with that task, we offer two valuable tools :

  • Tables of coordinate conversion between different species of coronaviruses, to help with the transfer of annotations
  • Tables of annotation of all residues. Compilated literature on Coronaviruses links each residue to their functional annotation, including from other species of coronavirus.

To illustrate the process, we studied a non-synonymous polymorphism within the S-gene found in the previous portion. We are trying to interpret its possible effect.

# Outline

Obtain coding sequences of S proteins from a diverse group of coronaviruses, and generate amino acid alignment to create a table of coordinate conversion.

# Input

Downloaded CDS sequences of coronavirus Spike proteins from NCBI Viral Resource for the following coronaviruses:

Accession Description
FJ588692.1 Bat SARS Coronavirus Rs806/2006
KR559017.1 Bat SARS-like coronavirus BatCoV/BB9904/BGR/2008
KC881007.1 Bat SARS-like coronavirus WIV1
KT357810.1 MERS coronavirus isolate Riyadh_1175/KSA/2014
KT357811.1 MERS coronavirus isolate Riyadh_1337/KSA/2014
KT357812.1 MERS coronavirus isolate Riyadh_1340/KSA/2014
KF811036.1 MERS coronavirus strain Tunisia-Qatar_2013
AB593383.1 Murine hepatitis virus
AF190406.1 Murine hepatitis virus strain TY
AY687355.1 SARS coronavirus A013
AY687356.1 SARS coronavirus A021
AY687361.1 SARS coronavirus B029
AY687365.1 SARS coronavirus C013
AY687368.1 SARS coronavirus C018
AY648300.1 SARS coronavirus HHS-2004
DQ412594.1 SARS coronavirus isolate CUHKtc10NP
DQ412596.1 SARS coronavirus isolate CUHKtc14NP
DQ412609.1 SARS coronavirus isolate CUHKtc32NP
MN996528.1 SARS-2 Cov
MN996527.1 SARS-2 Cov
NC_045512.2 SARS-2 Cov
NC_002306.3 Feline infectious peritonitis virus
NC_028806.1 Swine enteric coronavirus strain Italy/213306/2009
NC_038861.1 Transmissible gastroenteritis virus

These viruses were chosen based on a publication by Duquerroy et al. (2005). The sequences were extracted manually--a painful process.

# Output

We produce two alignments, one at the nucleotide and one at the amino acid level, of Betacoronavirus spike proteins. The alignments can be visualized with the Multiple Sequence Alignment Visualization in Galaxy :

Visualization of amino acid alignment in Galaxy

Alignments of Spike proteins
Nucleic Alignment of Spike proteins
A. CDS alignments
Proteic Alignment of Spike proteins
B. Protein alignment

# Workflow and history

The Galaxy history containing the latest analysis can be found here. The publicly accessible workflow can be downloaded and installed on any Galaxy instance. It contains all information about tool versions and parameters used in this analysis.

Analysis Workflow

The transeq tool converts the CDS sequences into protein sequences, which we then align with each other using mafft. The output is fed into tranalign along with the nucleotide sequences. tranalign produces a nucleotide alignment coherent with the protein alignment.

# Generation of Coordinate maps

We used this workflow to generate alignment across Coronaviruses for each gene of SARS-2 Cov. From these alignments, using mafft, we created coordinate conversion. The mafft -add option allows the addition of sequences to an alignment, but also generated mapping of the new sequence to the coordinate in the alignment. We reported these mapping data to the SARS-2 Cov coordinate for more clarity.

These tables can be queried through the notebook included in this section. To find positions equivalent to your residue or region of interest :

  • Select the Gene
  • Select the species in which your coordinates are
  • Select the region of interest

# Residue annotations

Since the beginning of this project, we have been compilating functional information from literature for each residue of each gene of SARS-2 cov. The literature covers several species of coronavirus, and we used the coordinate tables presented above to transfer the annotations between species. In cases where the residue is different from the one annotated, it is specified in the table, and all annotations are linked to their article of origin. Due to the considerable amount of literature available, the annotations are incomplete, but we are working on enriching them every day. You can contribute to the annotation effort by adding annotations in the tables on Github

# BioConda

Tools used in this analysis are also available from BioConda:

Name Link
mafft Anaconda-Server Badge
emboss Anaconda-Server Badge

# Notebooks

Source notebook: https://observablehq.com/@delphine-l/coordinate-conversion-between-species-of-coronavirus

Source notebook: https://observablehq.com/@delphine-l/annotation-of-sars-2-coronavirus-genome