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 - 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

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 = PYRIDAMINES

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.

Step 1  Unzipping the DNA Enzyme called helicase which breaks the hydrogen bonds between the base pairs. OPENS THE MOLECULE UP! Step 2  Complementary Base Pairing Each of the DNA strands can now act as templates to construct complimentary sides. Enzyme DNA polymerase attaches complementary nucleotides to each of the open strands

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

Not that simple though…… Antiparallel strands…. The two sides of the DNA molecule run in opposite directions. The 5 and 3 refer to the orientation of the ribose molecule

Because they are anti-parallel…. The copying can only happen in one direction along the 35 side of the template. Leading strand Built continuously as fork opens Lagging Strand Built in fragments, Needs patching together by ligase

Animations Overview http://highered.mcgraw- hill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::535::/sites/dl/free/0 072437316/120076/micro04.swf::DNA%20Replication%20Fork Detailed http://www.johnkyrk.com/DNAreplication.html

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

RNA vs. DNA 3 Main Differences Why use these replacements????? 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 Goal: Copy DNA code (gene) onto an mRNA molecule. Steps: Helicase opens DNA molecule at specific section (gene) Enzyme RNA polymerase binds to a promoter site (specific base sequence) just before the gene sequence RNA polymerase moves along gene sequence and creates a complementary RNA strand

Transcription Animation: http://www.stolaf.edu/people/giannini/flashanimat/molgenetics/transcription.swf

Transcription When RNA polymerase reaches terminator sequence (specific base sequence), new RNA molecule released from DNA molecule mRNA molecule travels to the cytoplasm where the message will be translated into polypeptides YOU TRY IT!

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!!!! Introns – What gets cut out (stay in the nucleus) Exons – What gets left it (Exits the nucleus) In prokaryotes, the mRNA is ready to go  No Editing Needed!

RNA EDITING Exons vs. Introns

Translation Goal: mRNA  Polypeptide (will be a protein once the 3D folding occurs) Review Proteins made by linking together amino acids 20 different amino acids Sequence of amino acids going to determine protein properties (shape). So how can a code of 4 letters (bases) code for 20 different amino acids?

The Genetic Code In the genetic code, three bases will code for one amino acid Why 3?  4^3 = 64 different combinations with nucleotide triplets, compared with 4^ 2 = 16 different combinations with pairs. Would 2 bases be enough??? The three bases that code for a specific amino acid are called codons ONE CODON = ONE AMINO ACID!!!!

Translation Codons- You are responsible for knowing 4: AUG UAA, UAG, UGA

Translation - Interpreting the Code Sequence of mRNA GGUACGUCCCCA Read as GGU-ACG-UCC-CCA

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 - Consists of two subunits

Translation in action….. Best to watch a video first Simple http://www.stolaf.edu/people/giannini/flashanimat/molgenetics/tr anslation.swf Complex http://highered.mcgraw-hill.com/olc/dl/120077/micro06.swf http://www.johnkyrk.com/DNAtranslation.html

Overall Process Initiation Start codon read, first tRNA binds to mRNA bringing methionine Ribosome assembles Elongation See animations for detail Chain builds Termination Stop codon reached Whole assembly falls apart Polypeptide formed

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 - 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

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

Mutations- So all mutations are bad right?????