DNA – Molecular Genetics

DNA STRUCTURE – Review…… -DNA is made of monomers called nucleotides. -Nucleotides consists of -5 carbon sugar -Phosphate group -Nitrogenous base (A,T,C,G) -Backbone- made of covalent bonds between bonds between the phosphate and sugar -“Rungs of Ladder” – made of pairs of nitrogenous bases.  A-T  C-G

Nitrogenous Bases  In DNA, there are 4 types of bases

STRUCTURE - -Sides of Ladder – Sugar/Phosphate backbone -Rungs of Ladder – Base Pairs -Two strands of DNA are connected through weaker Hydrogen bonds that form between bases - Only certain bases can form these hydrogen bonds with each other - They are called complementary base pairs

How many hydrogen bonds between C-G? A-T? Would A-G be possible????

CHARGAFF’S RULE  Amount of Adenine = The amount of Thymine  Amount of Guanine = The amount of Cytosine  If a sample of DNA contained 16% adenine……….?  PURINES = PYRIMIDINES

What does DNA need to be able to do?  Be copied– every time a cell divides  DNA Replication  Be “read” – every time we need to make a protein.  Transcription

DNA REPLICATION  Overview  Semi-conservative  Molecule opens up  Each side is a template to build a complementary side  End up with two DNA molecules, each one half old and half new.

Replication Before replication…. Think of DNA as a Closed zipper. A G T C A G T C Hydrogen bonds Covalent bonds

Replication Step 1: “Unzip the Zipper” Helicase breaks Hydrogen Bonds between chains - Opens the zipper A G T C A G T C

Replication Step 2: Each chain of nucleotides is used to build a two new chains using the enzyme DNA polymerase A G T C A G T C A G T C A G T C

Replication Step 2: Still must follow complementary base pair rules A binds with T G binds with C A G T C A G T C A G U C A G T C

PRACTICE

Semi-Conservative Replication -Each of the resulting DNA molecules are identical to each other and each contains one original strand and one new one.

Animations  Overview hill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::535::/sites/dl/free/ /120076/micro04.swf::DNA%20Replication%20Fork  Detailed 

Replication Bubbles  DNA is LOOOOOOOOONNNNNNGGGGGG!  In order to speed-up copying, replicate in several places at once.

RNA vs. DNA  3 Main Differences  Single Stranded  Sugar in RNA is ribose  In RNA, thymine is replaced by uracil  Why use these replacements?????  CHEAPER!!!!

3 types of RNA  Messenger (mRNA)  carries the RNA copy of the DNA gene out of the nucleus to the ribosomes.  Ribosomal (rRNA)  make-up portion of the ribosome.  Transfer (tRNA)  bring individual amino acids to the ribosome for assembly into polypeptides.

2 key steps to making proteins Transcription -Copy DNA code on mRNA Translation Reading the mRNA molecule code and turning it into a polypeptide chain (linking together amino acids)

Transcription Animation: cription.swf cription.swf

 Step 1: Open DNA zipper  Just like replication we first need to separate the DNA strands Transcription A G T C A G T C

 Step 2: Make the RNA  Using same pairing rules copy one chain of DNA into RNA  A’s in the DNA will pair with U’s in the RNA!! Transcription

A G T C A G T C A G U C

 Step 3: Chains of DNA pair up again, RNA gets kicked out Transcription A G U C A G T C A G T C Now we have a temporary copy of one of our recipes!

 mRNA molecule  Made in the nucleus  Needs to get to the ribosome in the cytoplasm. Transcription

Difference in cell types?  In eukaryotes, mRNA transcript has to be modified before leaving the nucleus to be translated.  RNA EDITING!  SIGNIFIGANCE – One gene can code for many different proteins!!!!  In prokaryotes, the mRNA is ready to go

Translation Goal: mRNA  Polypeptide (unfolded protein)  Review  Proteins made by linking together amino acids  20 different amino acids  We need to be able to use the mRNA code to determine which amino acids to put together.

Genetic Code: Think of the genetic code as a “Nucleotide to Amino acid” dictionary A U G G A G C Codon 1Codon 2

Genetic Code: Every three bases of the mRNA is called a Codon A U G G A G C Codon 1Codon 2

Genetic Code: Each Codon translates to a different Amino Acid. - cell reads codons to make protein A U G G A G C Codon 1Codon 2

Genetic Code: AUG is always the first codon – means “Start here!” GGC means “now go get glycine” UAG means “Stop! You’re done!” A U G G A G C Codon 1Codon 2

Genetic Code: There are 20 different amino acids and 64 codons. So… Some codons “mean” the same amino acids (GGC and GGG) A U G G A G C Codon 1Codon 2

Translation - Interpreting the Code  Sequence of mRNA GGUACGUCCCCA  Read as GGU-ACG-UCC-CCA Amino Acid Sequence?

Translation (the details) The players mRNA – code for making polypeptide rRNA – ribosome - assembles polypeptide tRNA – transfers amino acids to ribosome for assembly  All RNA made by the NUCLEOLUS

tRNA and rRNA structure/function tRNA is a folded RNA strand - amino acid on one end - anti-codon on the other end rRNA makes up part of the ribosomes - ribosomes made of rRNA and protein

Translation in action…..  Best to watch a video first  Simple  anslation.swf anslation.swf  Complex  

Translation: Step 1: mRNA lines up to be read by ribosome A U G G A G C C A G G U U A

Translation: Step 2: tRNA’s with anticodons that are complementary to codons line up A U G G A G C C A G G U U A UAC AA1 CUC AA2

Translation: Step 3: Amino Acid 1 is linked to amino acid 2 with a peptide bond A U G G A G C C A G G U U A UAC AA1 CUG AA2

Translation: Step 4: empty tRNA leaves and Ribosome shifts to a new codon A U G G A G C C A G G U U A U A C AA1 CUG AA2 GGU AA3

Translation: Step 5: steps repeat until stop codon is reached A U G G A G C C A G G U U A AA1 CUC AA2 GGU AA3 CCA AA4

Translation: Step 5: steps repeat until stop is reached U G G A G C C A G G U U A AA1 AA2 G G U AA3 CCA AA4 G A U C STOP

Translation: Step 6: protein is released U G G A G C C A G G U U A CCA AA1 AA2AA3 AA4 G AUC STOP

When things go wrong……  Review –  A protein’s shape determines its function  The sequence of amino acids determines its shape  DNA determines the sequence of amino acids..  WHAT IF WE CHANGE THE DNA????

Mutations  Mistakes happen!  Sometimes during DNA replication, bases can get inserted, removed or switched!!  Changes in the DNA base sequence are called MUTATIONS !!!  Can be single nucleotides – Point  Can be whole chunks of DNA- Chromosomal

Point Mutations  One base in the sequence is affected Substitution – one base switched out for another Insertion – base put into code Deletion – base removed from code

Point - Substitution  If this happens on a gene portion of the DNA molecule, it can result in a change in one of the amino acids in a polypeptide sequence.

Point - Insertion/Deletion  When a base is inserted or deleted, much bigger changes  Changes like this are called frame shift mutations  shift the reading frame of the genetic code. (how we read it)  These mutation affect all amino acids that follow the mutation point

Example of frameshift

Point - FRAMESHIFTS – class example  THE MAN SAW THE CAT HIT THE DOG Remember, frameshifts only involve insertion and deletions!

Point - Substitutions  Missense mutations  new nucleotide alters the codon so as to produce an altered amino acid in the protein product.  Can cause big changes in polypeptide if new amino acid in chain is chemically different than the one it replaced.  Can be neutral if new amino acid is chemically similar to one it replaced.

Point - Substitutions (cont.)  Nonsense mutations  With a nonsense mutation, the new nucleotide changes a codon that specified an amino acid to one of the STOP codons ( TAA, TAG, or TGA ).  Significance?  The earlier in the gene that this occurs, the more truncated (shortened) the protein product and the more likely that it will be unable to function.

Point - Silent mutations  Most amino acids are encoded by several different codons.  For example, if the third base in the TCT codon for serine is changed to any one of the other three bases, serine will still be encoded. Such mutations are said to be silent because they cause no change in their product and cannot be detected without sequencing the gene (or its mRNA).

Chromosomal Mutations  Involve large-scale changes to structure or number of a chromosome!  Have much greater consequences than point mutations  WHY???  MANY GENES AFFECTED!!!!