Protein Synthesis

Protein Synthesis Transcription and Translation AP Biology Unit 2

Flow of Genetic Information • All living organisms use DNA to synthesize RNA to make proteins • Same two-step process: Transcription  Translation • Some antibiotics inhibit protein synthesis in bacteria. • Ex. Neomycin (the antibiotic in Neosporin) interferes with the process of translation)

Genes and Chromosomes • DNA is organized into chromosomes • Humans have 46 chromosomes in each cell. • Genes are “coding” regions of DNA • Each gene is the code for how to make a specific protein. • Human chromosomes are made up of • DNA • Histone proteins that DNA is wound around

phosphate Sugar Base Structure of DNA • The carbons in the 5C sugar each have a number • Start to the right of the oxygen and go around clockwise 5 1 • This gives the nucleotide 2 distinct ends • 5’ end (closer to carbon 5) • 3’ end (closer to carbon 3) 4 3 2

A way to remember it: Human Nucleotide  Phosphate Base 5 C 4 C C 1 3C C 2

Nucleic Acid Structure • DNA is double stranded • Hydrogen bonds between bases • A pairs with T • C pairs with G • The strands are antiparallel • One strand runs 5’-3’ • The other runs 3’-5’ Image taken without permission from http://bcs.whfreeman.com/thelifewire

Question… • Why can’t the DNA strands be parallel (both running 5’-3’)? • This wouldn’t allow the bases to be near each other to hydrogen bond.

Transcription • DNA is transcribed into 3 kinds of RNA • mRNA = messenger RNA (the RNA code used to make protein) • tRNA = transfer RNA (participates in translation) • rRNA = ribosomal RNA (part of ribosomes) • RNA Polymerase is the enzyme that transcribes the DNA into RNA

Initiation • How transcription starts • RNA Polymerase recognizes a promoter sequence on the DNA • RNA Polymerase binds to the promoter • DNA is unwound to start transcription • What kind of bonds are being broken to unwind/separate the strands of DNA? • Hydrogen bonds

5’ 3’ 5’ 3’ Promoter Sequences • In prokaryotes, RNA Polymerase must find these sequences: • + 1 is the first base in the RNA (where the actual transcription of DNA starts from)

Eukaryotic Promoter Sequences • In eukaryotes, the RNA polymerase must find the following sequences: • Eukaryotic genes can also have enhancer sequences to help RNA polymerase bind • We’ll talk about these a little later– don’t worry about them right now 

What do you think this diagram shows about transcription? Bases changed to…

Promoters • In order for RNA Polymerase to recognize it, the promoter sequences • Must be the correct sequence of bases (small changes OK) • Must be correctly spaced apart • If these conditions aren’t met, RNA Polymerase can’t bind to the DNA and no transcription occurs.

Elongation • How the RNA strand is built • RNA Polymerase matches the appropriate (complementary) nucleotides to the DNA template strand • Template strand = the actual strand RNA Polymerase uses to build RNA • Coding (Nontemplate) strand = not used for building RNA, but has the same sequence as the RNA.

5’ 5’ 5’ 3’ 5’ 3’ 3’ 3’ Building the RNA • The RNA Polymer grows in a 5’-3’ direction • RNA Polymerase only adds new nucleotides on to the 3’ end. • Considering this, in what direction must the template strand of DNA be running? • 3’-5’ (since it is building its complement)

Question … • In terms of the sequence, how will the RNA differ from the sequence of the coding strand in the DNA? • T’s are replaced with U’s

Termination • How transcription of RNA ends • RNA Polymerase recognizes a termination signal on the DNA template • Usually a long string of A’s or a series of A’s and T’s • RNA Polymerase falls off the DNA template • Stability of mRNA is minutes  hours (depends on type of cell and RNA)

Question… • How do the specific chemical properties of the termination sequence cause termination to occur? • There are only 2 hydrogen bonds between A and T/U • With a string of A’s and U’s, there are much fewer bonds to hold the DNA template and RNA together  they separate  transcription ends

Translation • Using the mRNA code to create the appropriate protein. • Occurs in the cytoplasm/on the rough ER • Sequence of 3 nucleotides codes for a particular amino acid = codon • 64 different codons

Question… • Why can’t 1 or 2 nucleotides code for an amino acid? • Not enough combinations to code for all 20 amino acids • With 1 nucleotide  only 4 possibilities (A, C, G, U) • With 2 nucleotides  only 4 x 4 = 16 possibilities (AA, AU, AC, AG, CC, …)

The codon table

Amino acid attached here tRNA • tRNA brings the correct amino acid to match with the mRNA codon • Each tRNA holds a specific amino acid and has a particular anticodon. • Aminoacyl tRNA synthetases are enzymes that attach the correct amino acids to the tRNA

Question… • For the anticodon shown in the diagram, what would the complementary codon on the mRNA be? • 5’ UUC 3’ • Which amino acid is attached to this tRNA? • Phe

Ribosomes • Made up of 2 subunits • Composed of rRNA and protein • Not specific to any particular protein– can be used to translate any RNA into protein • Workbench for translation – holds mRNAs and tRNAs in the correct positions to assemble protein.

Ribosomes • 3 sites on the ribosome • A site = where tRNA first binds to mRNA • P site = where the amino acid is added on to the polypeptide chain • E site = exit site

Translation • Begins with the Start codon = AUG • Codes for methionine (Met) • Not the same thing as +1

Translation • Ribosome moves along mRNA in a 5’->3’ direction, catalyzing the translation of the mRNA into protein • breaks bond between tRNA and amino acid • creates a new peptide bond to link it to polypeptide chain

Question… • How does the mRNA know if it is correctly matched to the tRNA? • Hydrogen bonding between the bases is correct

Stopping Translation • Ribosome is released when a stop codon is reached • UAA, UAG, UGA = stop codons (don’t code for any tRNA anticodons) • A release factor binds to the mRNA instead • Ribosome breaks apart, mRNA and protein are released

Summary of Protein Synthesis • In Eukaryotes • How is it different in prokaryotes?

Why is this important? 1. Changes in the DNA sequence will lead to changes in the transcribed _________. 2. This results in a different codon which may code for a different ______________. 3. A different ___________ means a different R group. 4. A different R group may have different chemical properties. 5. These different chemical properties may lead to a different protein ____________. 6. A different protein structure may affect its _________! 7. See how this is all starting to connect! Exciting!!! 

Why is this important? 1. Changes in the DNA sequence will lead to changes in the transcribedRNA. 2. This results in a different codon which may code for a different amino acid. 3. A different amino acidmeans a different R group. 4. A different R group may have different chemical properties. 5. These different chemical properties may lead to a different protein structure. 6. A different protein structure may affect its function! 7. See how this is all starting to connect! Exciting!!! 

microRNAs and RNAi • Small, single stranded RNA molecules (miRNAs and siRNAs) • microRNA = miRNA • Small interfering RNA = siRNA • Bind to complementary sequences in mRNA molecules • Can control the expression of (translation of) specific RNA molecules

Question… • How will microRNAs disrupt translation? • Block translation by creating a physical road block • RNA-RNA binding also marks the mRNA for degradation • Called RNAi = RNA interference

Protein Synthesis