Download determining evolutionary relationships using

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DETERMINING EVOLUTIONARY RELATIONSHIPS
USING PROTEINS
Introduction:
In this course you have learned that living organisms regardless of species are united by
one common element…….we all have hereditary material in our cells which holds the instructions
for making protein. The cells of all organisms recognize the language of DNA and thus are capable
of producing protein from DNA instructions. Genes hold the instructions for the proteins that
each living organism makes and those proteins determine the traits that an organism has.
Different organisms have different types of genes for their different traits. Genes can also be
shared amongst organisms of different species, though. For example, the gene that makes
hemoglobin protein is present in many living organisms of different species. This unit has you
investigating how organisms change over time through Evolution. You know that organisms in a
population have variation in traits caused by mutation to the DNA code. Some of those variations
end up being favorable to an organism's survival because of the environment in which they live. It
is through this mechanism (natural selection) that a species changes over time.
While DNA is responsible for coding for specific proteins, to some degree, all organisms
share similar proteins. These similar proteins are inherited from a common ancestor. When a cell
divides, random errors in DNA nucleotide sequences occur. These errors may lead to the
emergence of different proteins in different organisms. Therefore, the more differences in
protein biology two organisms share, the more time has elapsed between the two organisms'
divergence from a common ancestor. We would also expect two organisms that share a high level
of protein similarity to have similar anatomical features and be closely linked phylogenetically.
Using the process of electrophoresis, scientists can easily isolate molecular fragments, the most
common being nucleic acids (DNA and RNA) and proteins, from living material. In protein
electrophoresis, fragments of proteins are separated on a gel by the size of the molecules, and
they appear on the gel as bands. The larger the bands, the more of a certain size protein is
present in the sample. Two samples that appear to have similar bands likely share similar proteins
and probably are closely linked evolutionarily. By separating protein molecules, scientists can
determine which organism share the most identical material and theorize about lineages of
different species.
Lastly, scientists have also been able to determine the amino acid sequences of specific
proteins in particular organisms. After determining amino acid sequences and DNA sequences,
scientists publish their findings on public databases. These published molecular sequences can be
accessed on bioinformatics websites and used to determine evolutionary relationships of
organisms.
BIOINFORMATICS AND DETERMINING
EVOLUTIONARY RELATIONSHIPS
In this part of the lab you will learn how to use bioinformatics websites in order to
help you deduce evolutionary relationships amongst organisms. As mentioned previously,
two organisms that share a high degree of protein similarity are more closely related in
evolutionary terms than two organisms who have many differences in their proteins. You
will use the Biology Workbench in order to determine the evolutionary relationships
between the following organisms: http://workbench.sdsc.edu/
HUMAN
GORILLA
GIBBON
CHIMP
HORSE
IGUANA
1) You have to set up a free account with the Biology Workbench. Paste link above
into a new browser and click "register for a free account." You will have to put a
valid email address in to register. RECORD YOUR LOGIN AND PASSWORD
BELOW!!
LOGIN:
PASSWORD:
2) Click on the graphic for the Biology Workbench shown at the top of the page once
entering the database to start your work!!
3) Before starting you session you will name your work. Scroll down and click on the
button for "session tools." Next you will click on the pull down menu and select "start
a new session," click "run." Name your session by typing "hemoglobin and evolution"
into the session description box. Finish by clicking on the "start a new session"
button.
OBTAINING THE AMINO ACID SEQUENCES!
4) Now you're ready to begin using the workbench!! Click on the button for "Protein
tools." You are searching for amino acid sequences, so you use protein tools. If you
were looking for DNA sequences, what button would you click on???????
5) Using the pull down menu again, select the function "Ndjinn" for a multiple database
search. Scroll down the page and find the database named, "Swissprot." Click on the
box next to Swissprot!
6) Now you're all set to search for an amino acid sequence. In the search box type the
phrase, "Human Beta Hemoglobin." Then click on the "search" button.
7) The database will reveal a number of search results. Scroll through the choices and
only select the one which displays the sequence for the "Homo Sapien Beta subunit
of Hemoglobin protein." Once you have found this sequence, click on the "import
sequence" button in order to save your amino acid sequence.
8) You will now need to repeat steps 5 through 7 in order to obtain the hemoglobin
protein sequences for the gorilla, gibbon, chimp, horse, and iguana.
ANALYZING THE SEQUENCES FOR SIMILARITIES AND DIFFERENCES!
9) Once you have all the amino acid sequences for the organisms listed above, you're
now ready to compare the sequences. The comparison will show you the differences
in amino acid sequence for the six organisms' beta hemoglobin protein. To do this
click on all six of the boxes next to each organism's protein sequence. Once
you've click on all six boxes, use the pull down menu in order to select the
"CLUSTALW" function for a multiple sequence alignment. On the next page you come
to, select the "submit" button.
10) Now you will have the alignment of all six amino sequences for hemoglobin protein.
Paste the alignment in the text box below. How do you know where the sequences
differ?? Answer this question below. Hint…..color has a lot to do with it. Remember
that each letter represents 1 amino acid!!
11) To save your alignment of the amino acid sequences click on the "import alignment"
button. This will now automatically take you to the "alignment tools" section of the
data base. You will now construct a phylogenetic tree based upon the hemoglobin
protein comparison for these six organisms.
CREATING A PHYLOGENETIC TREE!
12) To draw a phylogenetic tree you should first click on the box next to the saved amino
acid sequence alignment. Once you've done this, you can now use the pull down menu in
order to select the "DRAWGRAM" function. On the next page you will click on the
"submit" button. Paste your phylogenetic tree in the box below and use it to help you
answer the analysis questions which follow. Please label each of the abbreviations on the
tree with the organism's name that they represent. For example, gorgo = gorilla.
ANALYZING YOUR RESULTS:
1)
Using the phylogenetic tree you obtained, answer the questions which follow:
a. Which organisms had hemoglobin most like the human?
b. What does a very similar protein sequence suggest about two organisms?
c. Which organisms had hemoglobin least like the human?
d. What does a very different protein sequence suggest about two organisms?
2)
The six organisms you investigated in this lab all have hemoglobin protein. They have
differences in their versions of the protein, but they all still have hemoglobin
protein. What does this suggest about the protein, hemoglobin, in terms of its
importance to life?
3)
You've heard a little bit about taxonomy from your instructor. Look at the taxonomic
classification for the six organisms you investigated in this lab shown below.
CLASS
ORDER
FAMILY
GENUS
Human
Mammal
Primate
Hominidae
Homo
Chimpanzee
Mammal
Primate
Hominidae
Pan
Gorilla
Mammal
Primate
Hominidae
Gorilla
Gibbon
Mammal
Primate
Hylobatidae
Hylobates
Horse
Mammal
Laurasiatheira
Equidae
Equs
Iguana
Retile
Iguannia
Iguanidae
Iguana
a) To what extent do the taxonomic classifications shown above correlate with
what you found comparing proteins for the six organisms?
b) What can taxonomy tell us about evolutionary relationships?