Welcome to Introduction to Bioinformatics Wednesday, 25 January Introduction to Molecular Biology...

Welcome toIntroduction to Bioinformatics

Wednesday, 25 JanuaryIntroduction to Molecular Biology

Part 2: DNA to protein

• Coming attractions!• Significance• Palindromes (SQ4)• Why introns? (SQ8)• Types of mutation (SQ12)

Study Question 8Why do introns exist?

I do not understand why introns would even exist.

I dont understand why the information for a

single element would be spread out.

Well, they do seem pretty mysterious. I’ll quickly review what introns and

exons are and then propose an explanation as to why they might

serve a useful purpose.


DNA

Primary transcript(hnRNA)

Transcription

Transcription is a key step in the expression of genes, i.e. the chain of events that convert information

on DNA into cell behavior.

The end product of transcription is a molecule of RNA corresponding

to a small fraction of the DNA.


AAAAAA...AAA

DNA

Primary transcript(hnRNA) MeG cap Poly-A tail

Transcription

In eukaryotes, the primary transcript (i.e. the direct RNA product of transcription) is processed before it is used in translation.

One modification is a special guanosine placed at the 5’ end of the message.

Another is a string of adenosines placed at the 3’ end of the message.


AAAAAA...AAA

DNA


Transcription

They function in the stability of the message and in helping ribosomes bind to the RNA’s 5’ end.

However, poly-A tails are much less important in bacteria and their viruses, and caps are (to my knowledge) absent.

Since we will be focusing on bacterial viruses, these structures needn’t concern us now.


AAAAAA...AAA

DNA


Transcription

Transcripts are subject to a third modification. If you looked at the sequence of a transcript, you would not find anything in one region

to distinguish it from another.

However protein-nucleic acid complexes in the nucleus of eukaryotic cells recognize

certain sites ( ) in the message.


AAAAAA...AAA

AAAAAA...AAA

Splicing


Processedtranscript(mRNA)

MeG cap Poly-A tail

Intron

Exon

These complexes cause some of the RNA to be cut out, a process called splicing.

RNA that’s cut out is called an intron. RNA that’s left in is called an exon.


AAAAAA...AAA

AAAAAA...AAA

Splicing



MeG cap Poly-A tail

Intron

Exon

It is the mature, processed transcript that enters the cytoplasm and is translated by ribosomes.


AAAAAA...AAA

AAAAAA...AAA

Splicing



MeG cap Poly-A tail

Intron

Exon

Where are introns when translation takes place?

…what role [does] an

intron plays in translation?


AAAAAA...AAA

AAAAAA...AAA

Splicing



MeG cap Poly-A tail

Intron

Exon

You could answer this question the same way you discovered

what is the beginning of a gene.

What would you need to do this?

…could you explain… how a cell's apparatus

distinguishes exon from intron?


That explains what introns are, but not why they exist.

They’re not necessary… bacteria live perfectly well even though introns are very rare in bacteria and their viruses.

In eukaryotes, however, they are very common.

Why?

How do introns contribute to differences between eukaryotes and prokaryotes?


Huge differences amongst bacterial DNA sequences

Small differences in morphology

Bacterial sequences change rapidly in evolutionary time, but major changes in morphology are rare.


Small differences amongst vertebrate

DNA sequences

Large differences in morphology

http://uncyclopedia.wikia.com/wiki/User:Skinfan13/Upsilon_Sigma_Sigma/Collaboration/May2010collab

In contrast, eukaryotes can look very different, even though the sequences of their genes are similar.

The gene sequences of mammals are generally very similar to each other. They differ in the adjacent

regulatory sequences that control gene expression.


Hormone-responsive protein kinase

hormoneresponsiveness

protein kinase

Here’s an example of how introns can contribute

to a drastic change in phenotype without a drastic

change in sequence.

Suppose that there is a gene that encodes an enzyme that

responds to a hormone.

The first part of the gene encodes the part of the protein that

recognizes the hormone, and the rest encodes the enzyme.


Hormone-responsive protein kinase


DNA bindingprotein kinase

DNA-binding protein

Now suppose that there is a second gene, lying very far away on the chromosome (or even on a different

chromosome). This gene encodes a DNA-binding protein..




Since DNA frequently recombines, breaking and joining with other regions of the chromosome, the two genes may by

accident find themselves close to each other.

Maybe they’re close enough so that a single RNA is transcribed




Now the splice sites are close enough to produce, after splicing, an mRNA that would encode a novel protein.

In bacteria, the DNA recombination would have to be exact. In eukaryotes, somewhere in the vicinity will do.

Hormone-responsive DNA-binding protein




Introns therefore can greatly accelerate the rate of evolution of proteins of novel functions.

For more on exon shuffling, see Keren H et al (2010). Nature Rev Genet 11:345-355.

Hormone-responsive DNA-binding protein

Welcome toIntroduction to Bioinformatics

Wednesday, 25 JanuaryIntroduction to Molecular Biology

Part 2: DNA to protein

• Coming attractions!• Significance• Palindromes (SQ4)• Why introns? (SQ8)• Types of mutation (SQ12)

Study Question 12Single mutation from AGA

If this question is confusing, take it apart bit by bit.


…the AGA codon… That, at least, might be clear.

Write it down!

AGA arginine


…a single basepair mutation… What could that mean?

AGA arginine


Basepair? Of course DNA is double stranded, so a codon in DNA is paired

with complementary nucleotides.

What would be the basepairs of the AGA codon?

AGA arginine


Wait, that’s not enough. With double stranded DNA, you need to show

the direction of each strand.

Where should you put 5’ and where 3’?

AGA arginineTCT


Yes. Codons (like any other DNA or RNA sequence) are read 5’ to 3’.

What kind of single basepair mutations are possible?

What’s an example of a single basepair?

AGA arginineTCT

5’-3’-

-3’-5’


That’s one basepair.

What are some possible changes?

AGA arginineTCT

5’-3’-

-3’-5’


OK, that’s one possible change.

How many total possible changes are there?

Note I said “possible”. A G/A basepair is not possible.

AGA arginineTCT

5’-3’-

-3’-5’

GGA arginineCCT

5’-3’-

-3’-5’


I’m not going to write basepairs anymore – you can always imagine them.

Nor am I going to write 5’ and 3’. Remember, unless specified, left-to-right is always 5’ to 3’.

AGA arginine


Moving on, what could “encoding arginine” mean?

I’ll pause so you could look up what arginine is. (Google is your friend)

AGA arginine


…an amino acid.

Codons encode amino acids.

AGA arginine


So putting it all together….

…what does the first part of the question mean?

AGA arginine


Single basepair mutations in AGA change the codon.

Changing the codon may change the amino acid that’s encoded.

Which single basepair changes produce which changes in encoded amino acids?

AGA arginine


What might these terms mean?

They’re likely to refer to the result of the changes described in the first part of the question, changes in codons and perhaps

the amino acids they encode.

Start with two we’ve discussed: hydrophobic and hydrophilic

AGA arginine


Look at Figure 2 from the notes on Proteins. Notice that the amino

acids are arranged in a very similar manner to the generic amino acid

shown in Figure 1.

There is a constant part that all amino acids have, and there

is a variable part, labeled R.

Proteins, Figure 2 Proteins, Figure 1

Derived from Elseth & Baumgardner, Principles of Modern Genetics (1995), West Pub


I’ve represented the constant part as shadowed.

What distinguishes one amino acid from the others is what’s left.

Consider the amino acids with hydrophobic R groups. What generality can you draw from their chemical constituents?




Look at them!

Except for methionine, they consist solely of carbons and hydrogens.

They’re hydrocarbons, like oil.

What about the negatively charged R groups?






All the negatively charged R groups have free

carboxylate moieties ( ), with negative charges.

Positively-charged R groups?

-C-O-

O




All positively charged nitrogen atoms in some form.

Finally, the hydrophilic, uncharged R groups?




…a mixed bag, but all of them have uncharged polar groups,

for example -OH.

Of course, they are different in other ways: some are large,

some small, some have aromatic rings, others not.

Each has a different flavor.


Now back to the question.

AGA encodes arginine.Mutations can change the codon.

That might change the encoded amino acid.That might change the type of amino acid.

We have something that describes the types of amino acids (left). We need a table that

connects amino acids to their codons.

AGA arginineGGA ???


Here's such a table.

You can find dozens of these on the web, or use the genetic table found on the Resources & Links section.

First of all, does AGA indeed

encode arginine?

Learn how to use this table.


How many ways can AGA be

changed by a single nucleotide?

Answer this by counting, and then figure out how you

could calculate the number.

How many ways could a 10-letter

sequence be changed by a single

nucleotide?


…Back to AGA.

Categorize each change to AGA, using the labels

given in the question. How does the encoded amino acid change in each

case?

• If "conservative" is "close enough", how close is that?

Study Question 12Single mutation from AGA(Frequently asked questions)

A conservative amino acid substitution is one that should not affect the function of the

protein. However, this is often not easy to predict. In one context, leucine and

isoleucine might be interchangeable, while in another, substituting isoleucine for leucine

could lead to an inactive protein.

That isn't a satisfying answer, but it does reflect our current state of knowledge.



If you do, you will.

If the time ever comes when you find yourself constantly going back to a list

of amino acids and their properties, you will memorize them without trying.

Until then, the list is always available.

• Do we have to learn the 20 amino acids?



Well now, funny you should ask!

• Do we have to learn the 20 amino acids?

• Will we be able to decipher the genetic code of an alien being?

If you go to the Intro to Molecular Biology topic page, you will find a link to

an investigation called Alien Genetic Code.

Alien Genetic Code (Coming Attractions!)

CCCCCCCCCCCC Pro-Pro-Pro-Pro…

Terrestrial Genetic Code

Terrestrial ribosomes

There you will be given the means to perform virtual experiments.

Using terrestrial cell extracts and the RNA CCCCC…, these experiments would produce poly-proline, according to our genetic code.

CCCCCCCCCCCC

Alien Genetic Code

Alien ribosomes

? ? ?

`

Alien Genetic Code (Coming Attractions!)

But using alien cell extracts... who knows?

You can know.

Welcome to Introduction to Bioinformatics Wednesday, 25 January Introduction to Molecular Biology...

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