List of content:

# Topics Page no.

1 Introduction 4-5
2 Tools 6
3 Protein Sample Sequences 6
4 Methods and Materials 7-15
5 Discussion 16
6 Conclusion 16-17
7 References 17
1. Introduction
Coronaviruses are species of virus belonging to the subfamily Coronavirinae in the family
Coronaviridae, in the order Nidovirales. Coronaviruses are enveloped viruses with a positive-
sense single-stranded RNA genome and with a nucleocapsid of helical symmetry. The
genomic size of coronaviruses ranges from approximately 26 to 32 kilobases, the largest for
an RNA virus (1).

The name "coronavirus" is derived from the Latin corona, meaning crown or halo, and refers
to the characteristic appearance of virions under electron microscopy (E.M.) with a fringe of
large, bulbous surface projections creating an image reminiscent of a royal crown or of the
solar corona. This morphology is created by the viral spike (S) peplomers, which are proteins
that populate the surface of the virus and determine host tropism.

Proteins that contribute to the overall structure of all coronaviruses are the spike (S),
envelope (E), membrane (M) and nucleocapsid (N). In the specific case of the SARS
coronavirus, a defined receptor-binding domain on S mediates the attachment of the virus to
its cellular receptor, angiotensin-converting enzyme 2 (ACE2). Some coronaviruses
(specifically the members of Betacoronavirus subgroup A) also have a shorter spike-like
protein called hemagglutinin esterase (HE) (2)

Taxonomy (3):

1. Genus: Alphacoronavirus

Species: Alpaca coronavirus, Alphacoronavirus 1, Human coronavirus

229E, Human Coronavirus NL63, Miniopterus Bat coronavirus 1,
Miniopterus Bat coronavirus HKU8, Porcine epidemic diarrhea virus,
Rhinolophus Bat coronavirus HKU2, Scotophilus Bat coronavirus 512.

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2. Genus: Betacoronavirus

Species: Betacoronavirus 1, Human coronavirus HKU1, Murine

coronavirus, Pipistrellus Bat coronavirus HKU5, Rousettus Bat coronavirus
HKU9, Severe acute respiratory syndrome-related coronavirus, Tylonycteris
Bat coronavirus HKU4, MERS-CoV, Human coronavirus OC43, Hedgehog
coronovirus (EriCoV)

3. Genus: Gammacoronavirus

Species: Avian coronavirus, Beluga whale coronavirus SW1, Duck

coronavirus, and Infectious bronchitis virus

4. Genus: Deltacoronavirus

Species: Bulbul coronavirus, Munia coronavirus HKU13, Thrush

coronavirus HKU12.

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2. Tools
i. National Center for Biotechnology Information (NCBI)
ii. Gedit (software to open FASTA files)
iii. Clustal Omega (for multiple sequence alignment)

3. Protein Sample Sequences

i. Alphacoronavirus - Alpaca coronavirus

 AOC31982.1 S1
 AOC31981.1 S1
ii. Betacoronavirus - Human coronavirus
 AKL87245.1
 AKL87244.1
iii. Gammacoronavirus - Avian coronavirus
 ARJ35798.1
 ARJ35795.1
iv. Deltacoronavirus - Bulbul coronavirus
 ACJ12048.1
 ACJ12046.1

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4. Methods and Materials
i. Acquiring protein sequences from National Center for Biotechnology Information’s
(NCBI) database:
 Step 1 – Go the NCBI web site.

 Step 2 – Open the protein database.

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 Step 3 – Search for the desired protein sequences.

 Step 4 – Download the sequences in the FASTA format.

[For all the four species]

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  Step 5 - Open the sequences with “Gedit”.
 Results
Alpaca coronavirus

Human coronavirus

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Avian coronavirus

Bulbul coronavirus

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ii. Align all the sequences with the help of Clustal Omega (online)
 Step 1 – Go to the Clustal Omega web site.

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 Step 2 – Load the downloaded sequences on to the site

 Step 3 – Complete the following steps.

 Step 4 – Check email and follow the link

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 Step 5 – Select “show colour” for a better visual representation of the data.

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iii. View the phylogenetic tree.
 First the cladogram shows us the branches from a common ancestor.

 Selecting the “real” shows us the actual phylogenetic tree, which accurately portrays
the time line from when each of the branching took place from the common ancestor.

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5. Discussion
We were able to successfully obtain an accurate phylogenetic tree using Clustal Omega as all
the sequences were on similar lengths and so a global alignment technique was necessary.

However, due to the version being an updated one, when we typed “PROTEIN” in the Clustal
program (step 1) it automatically selected the BLOSUM 62 (BLOck SUbstitution Matrix) as
its default algorithm. Which in previous versions would have to be manually selected.

6. Conclusion

It can be seen that the sequences AOC31982.1 and AOC31981 (both from the specie Alpaca

coronavirus) and the sequences AKL87245.1 and AKL87244.1 (both from the
specie Human coronavirus) are closely related and are the most recent to have

undergone a mutation (evolution). The sequences ARJ35798.1 (from the specie

Avian coronavirus) and ACJ12048.1 (from the specie Bulbul coronavirus) are
closely related and had undergone some mutation much earlier on the biological

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timeline. Similar is the case with the sequences ARJ35795.1 (from the specie
Avian coronavirus) and ACJ12046.1 (from the specie Bulbul coronavirus).

But one thing can be ascertained from this data, and that is that all these species
of Coronaviruses share a common ancestry.

7. References:
1. Groot RJ, Baker SC, Baric R, Enjuanes L, Gorbalenya AE, Holmes KV, Perlman S,
Poon L, Rottier PJ, Talbot PJ, Woo PC, Ziebuhr J. "Family Coronaviridae". In AMQ
King, E Lefkowitz, MJ Adams, and EB Carstens. Ninth Report of the International
Committee on Taxonomy of Viruses. Elsevier, Oxford, 2011. pp. 806–828.
2. Li F, Li W, Farzan M, Harrison SC. "Structure of SARS coronavirus spike receptor-
binding domain complexed with receptor", 2005. Science. 309 (5742): 1864–8.
3. International Committee on Taxonomy of Viruses, 2010.

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