Watson and Crick In 1800’s it was believed that protein carried genetic information. By late 1940s it was accepted that DNA was the genetic material. Discovered.

Watson and Crick

• In 1800’s it was believed that protein carried genetic information.

• By late 1940s it was accepted that DNA was the genetic material.

• Discovered the structure for DNA in 1953– DNA made of nucleotides– Bases: A, T, C, and G– Used work of several other scientist to come up

with the structure of DNA

Rosalind Franklin

• Used X-ray diffraction to study the structure of DNA– Technique uses x-rays of DNA. X-rays bounce off a

sample of material to determine characteristics of the sample such as physical structure and chemical composition.

– Watson and Crick attend her lecture in 1951 on the structure of DNA. They used her information that DNA is a twisted helix.

Wilkins

• Maurice Wilkins was also studying DNA. He showed Franklin’s work to one of his peers- Watson

• 1952, Chargaff had found that DNA contained equal amount of A and T and equal amounts of G and C. – helped with base pairing.

• 1962, Watson, Crick, and Wilkins shared the Nobel Prize in physiology and medicine

• What happened to Franklin’s contribution to this project?

Chargaff

• Studies the nitrogenous bases of DNA• He found that a cell always has an equal

amount of adenine and thymine• Also an equal amount of guanine and cytosine• Base pairing rules- A-T and C-G

Chapter 8Medicine from Milk

Copyright © 2012 by W. H. Freeman and Company

Medicine for Milk

• Art of “Pharming”• Using genetically engineered modified animals

to churn out therapeutic drugs• Transgenic animals• First drug produced by transgenic animals is

already available, human protein called antithrombin

• Extracted from transgenic goats’ milk

Antithrombin Deficiency

• Antithrombin is most commonly used to treat patients who either inherit or acquire a deficiency of the antithrombin protein

• Patient develops dangerous blood clots• Antithrombin protein use to be isolated from

human blood – small amounts• Transgenic goat can produce large amounts of

antithrombin protein• Why is goat better than human?

What is a protein?• A protein is a macromolecule made up of

repeating subunits known as amino acids.– Proteins have many functions: they

allow our muscles to contract, give our hair and skin its texture, and facilitate thousands of chemical reactions in our cells.

Amino acids• Amino acids are the building blocks of proteins. • There are 20 different amino acids.– All amino acids have the same basic core structure, but

each also has a unique chemical side group.

Sizes of proteins

• Human antithrombin is 432 amino acids– Many human proteins are in this size range

• Protein titin for muscle contraction is 34, 350 amino acids long- longest human protein

Amino acids determine the shape and function of a protein

• Amino acids bond together to form linear chains. The sequence of amino acids in any given chain makes the chain unique.

• The chain folds into a 3-D protein based on the sequence of amino acids.

Amino acids determine the shape and function of a protein

• Changing an amino acid in the sequence changes the 3-D shape of the protein.– The shape of

the protein determines its function.

– 801

Where do protein’s come from?

• Cells make proteins using building blocks from our diet.

• The instructions to make the protein are encoded in our DNA – our genes.

Chromosomes include gene sequences that code for proteins

• Chromosomes consist of many genes along their length.– A gene is a sequence of DNA that contains the instructions

to make at least one protein.

Chromosomes include gene sequences that code for proteins

• Genes are expressed when the cell uses the DNA instructions to make proteins.

• Gene expression• 804

Genotypes and phenotypes

• The genotype of an organism is the genetic make-up of that organism.– Genes provide our genotype, proteins specified by

these gene determine of physical traits.

• The phenotype is the physical attributes of the organism – Antithrombin gene is on chromosome 1 and holds

instructions to make 432amino acids of the protein.- Cells express the gene to make the protein

Antithrombin prevents blood clots

• Antithrombin is a protein that helps prevent blood clots (thrombosis).– Rare condition (1 in 5000)

– The antithrombin gene is expressed by cells in the liver, which then release antithrombin protein into the bloodstream.

– Inactivates enzymes that promote blood clotting

Antithrombin prevents blood clots

• When people inherit antithrombin deficiency, it means that both of their copies of the antithrombin gene are defective.– You inherit one copy of chromosome 1 from the

mother and one copy of chromosome 1 from your father.

– This can happen because there are alternate versions of genes.- differ slightly in the nucleotide sequence

Different alleles influence phenotype

• Alleles are alternative versions of the same gene that have different nucleotide sequences.

• 807•

Antithrombin Deficiency

• People with inherited antithrombin deficiency usually takes medication to thin their blood and prevent clots.

• It takes 50,000 donors to produce 1kg of antithrombin.

• One trangenic goat can produce the same amount in 1 year.

Genes have regulatory sequences and coding sequences

• Two parts of a gene:• Genes are organized into

two parts:– Regulatory sequences

determine when and how much protein a gene makes.

– Coding sequences determine the amino acid sequence of the encoded protein.

MakingTransgenic Goats

• All mammals produce proteins and secrete them into their milk

• Isolate a human gene and transfer into DNA of a large mammal, like goat (goat embryo)

• Use regulatory sequence of goat and human protein.

• Gene is expressed in mammary gland.• Goat produces human proteins in milk• Collect protein in goat milk

Making a transgenic goat• Transgenic organisms carry one or more genes from a

different species.– Transgenic organisms are also called genetically modified

organisms (GMOs) because they have been genetically altered by humans.

Making a transgenic goat• Step 1: Create hybrid gene.– Goat regulatory sequence and human antithrombin coding

sequences are cut out of donor cell chromosomes and joined together using special enzymes.

Making a transgenic goat

• Step 2: Microinjection and embryo transfer.

Making a transgenic goat• Step 3: Purify antithrombin from transgenic milk.– Antithrombin protein is expressed in the milk of transgenic

females. This protein can be isolated from the milk and used to treat antithrombin deficient people.

Gene therapy

• Gene therapy is a type of treatment that aims to cure disease by replacing defective genes with functional ones.

• Gene therapy is used to treat severe combined immunodeficiency syndrome (SCID)– Disorders in which babies are born with deficient

immune systems.

Genetically modified organisms serve many purposes

• Crops can be modified to contain genes for natural pesticides, which reduces the amount of pesticide a farmer must use.

• Transgenic animals can be used to research a gene’s function or to produce marketable products like medicine and fibers.

Gene expression: An overview

• Gene expression has two main steps: transcription and translation.

Where does gene expression occur?• Transcription occurs in the nucleus of eukaryotic cells. • Transcription occurs in the cytoplasm of prokaryotic cells. Why?• Translation occurs on ribosomes. Ribosomes are the cellular

machinery used for protein synthesis.

Gene expression: Transcription• Transcription copies the coding sequence of DNA into the

complementary messenger RNA (mRNA) sequence.

Gene expression: Transcription

• RNA polymerase (function) binds to the regulatory sequence just ahead of the gene’s coding region. The DNA strands unwind, exposing the coding sequence of the gene.

Gene expression: Transcription

• The RNA polymerase moves along the DNA strand, “reading” the DNA coding sequence and synthesizing a complementary mRNA strand.

Gene expression: Transcription

• The complementary mRNA forms from a DNA template according to the rules of base pairing, except that in RNA, adenine (A) pairs with uracil (U).

Gene expression: Transcription

• As the mRNA strand is formed, it detaches from the DNA sequence, and the DNA reforms its double-stranded helix.

Gene expression: Transcription

• Once the mRNA molecule is complete, it leaves the nucleus. The gene remains part of the chromosome in the nucleus where it can be used again for transcription.

Gene expression: Translation

• Translation uses the mRNA sequence to assemble the appropriate amino acid sequence of the protein.

Gene expression: Translation• The transcribed mRNA associates with a ribosome.• What is the function of the ribosome?

Gene expression: Translation

• The ribosome moves along the mRNA, “reading” it in groups of three nucleotides (codons).

• Each codon specifies a particular amino acid.

Gene expression: Translation

• Transfer RNA (tRNA) carries an amino acid to the mRNA by using its anticodon to find a matching mRNA codon.

Gene expression: Translation

• When the correct tRNA is in place, the specified amino acid is added to the growing chain, and the ribosome moves on to the next codon.

Gene expression: Translation

• The finished amino acid chain detaches from the ribosome and folds into its 3-D shape.

• Animation 809

The genetic code

• mRNA codons specify particular amino acids according to the universal genetic code.

Transcription/Translation Exercise

DNA A G C G C G C T T A C G T A G T A G

mRNA U C G C G C GA A UG CA U C A U C

What is the role of tRNA?_correct AA________

Protein SER-ARG-GlU-Cys-ISO-ISO

Predicting mRNA and amino acid sequences from DNA sequences

• It is possible to use the rules of base pairing and the genetic code to predict the amino acid sequence that will result from a particular DNA template sequence.