1. From Gene to Protein How Genes Work
SLIDE SHOW BY KIM FOGLIA (modified) All Blue edged slides are Kim’s (hyperlinks may have been added)
How Genes Work

2. Essential knowledge 3 .A.1: DNA, and in some cases RNA, is the primary source of5. DNA replication ensures continuity of hereditary information c. Genetic information flows from a sequence of nucleotides in a gene to a sequence of amino acids in a protein.
1. The enzyme RNA polymerase reads the DNA molecule in the 3’ to 5’ direction and synthesizes complementary mRNA molecules that determine the order of amino acids in the polypeptide.
2. in eukaryotic cells the mRNA transcript undergoes a series of enzyme-regulated modifications To foster student understanding of this concept, instructors can choose an illustrative example such as:
Addition of a poly-A tail.
Addition of a GTP cap
Excision of introns
3. Translation of the mRNA occurs in the cytoplasm on the ribosome In prokaryotic organisms, transcription is coupled to translation of the message Translation involves energy and many steps, including initiation, elongation, and termination X The details and names of the enzymes and factors involved in each of these steps are beyond the scope of the course and the AP Exam.
The salient features include: i. The mRNA interacts with the rRNA of the ribosome to initiate translation at the (start) codon ii. The sequence of nucleotides on the mRNA is read in triplets iii. Each codon encodes a specific amino acid, which can be deduced by using a genetic code chart. Many amino acids have more than one codon iv. tRNA brings the correct amino acid to the correct place on the mRNA v. The amino acid is transferred to the growing peptide chain vi. The process continues along the mRNA until a “stop” codon is reached vii. The process terminates by release of the newly synthesized peptide/protein. d. Phenotypes are determined through protein activities

3. Essential knowledge 4 .A.1: The subcomponents of biological molecules and their sequence determine the properties of that molecule. a. Structure and function of polymers are derived from the way their monomers are assembled. 1. in nucleic acids, biological information is encoded in sequences of nucleotide monomers. Each nucleotide has structural components: a five carbon sugar (deoxyribose or ribose), a phosphate and a nitrogen base (adenine, thymine, guanine, cytosine, or uracil). DNA and RANA differ in function and differ slightly in structure, and these structural differences account for the differing functions [See also 1.D.1, 2.A.3, 3.A.1] b. Directionality influences structure and function of the polymer. 1. Nucleic acids have ends, defined by the 3’ and 5’carbons of the sugar in the nucleotide, that determine the direction in which complementary nucleotides are added during DNA synthesis and the direction in which transcription occurs (from 5’ to 3’). [See also 3.A.1] 2. Proteins have an amino (NH3) end and a carboxyl (COOH) end, and consist of a linear sequence of amino acids connected by the formation of peptide bonds by dehydration synthesis between the amino and carboxyl groups of adjacent monomers. LO 4.1 The student is able to explain the connection between the sequence and the subcomponents of a biological polymer and its properties [See SP 7.1] LO 4.2 The student is able to refine representation and models to explain how the subcomponents of a biological polymer and their sequence determine the properties of that polymer [See SP 1.3] SP1. The student can use representation and models to communicate scientific phenomena and solve scientific problems. SP 7. The student is able to connect and relate knowledge across various scales, concepts and representations in and across domains.

4. DNA gets all the glory, but proteins do all the work!
The “Central Dogma”
Flow of genetic information in a cell
How do we move information from DNA to proteins?
transcription
translation
DNA
RNA
protein
trait
To get from the chemical language of DNA to the chemical language of proteins requires 2 major stages:
transcription and translation
DNA gets all the glory, but proteins do all the work!
replication

5. from DNA nucleic acid language to RNA nucleic acid language
Transcription
from DNA nucleic acid language to RNA nucleic acid language
Transcription animation

6. DNA RNA RNA ribose sugar N-bases single stranded lots of RNAs
uracil instead of thymine
U : A
C : G
single stranded
lots of RNAs
mRNA, tRNA, rRNA, siRNA…
transcription
DNA
RNA

7. 3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS
RIBOSOMAL RNA (rRNA)
Made in nucleolus
2 subunits (large & small)
Combine with proteins to form ribosomes
Bacterial ribosomes different size than eukaryotic ribosomes
Evidence for ENDOSYMBIOTIC THEORY
Medically significant-some antibiotics target bacterial ribosomes w/o harming host
rRNA and t-RNA images from Image from: Biology; Miller and Levine; Pearson Education publishing as Prentice Hall; 2006
mRNA image from

8. 3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS
TRANSFER RNA (tRNA)
ANTICODON sequence
matches CODON on mRNA
to add correct
amino acids during
protein synthesis
AMINOACYL-tRNA SYNTHETASE
Enzyme attaches a specific
amino acid using energy from ATP

9. 3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS
MESSENGER RNA (mRNA)
carries code from DNA to ribosomes

10. Transcription Making mRNA transcribed DNA strand = template strand
untranscribed DNA strand = coding strand
same sequence as RNA
synthesis of complementary RNA strand
transcription bubble
enzyme
RNA polymerase
coding strand 3 A G C A T C G T 5 A G A A A G T C T T C T C A T A C G
DNA T 3 C G T A A T 5 G G C A U C G U T 3 C
unwinding G T A G C A
rewinding
mRNA
RNA polymerase
template strand
build RNA 53 5

11. RNA polymerases 3 RNA polymerase enzymes RNA polymerase 1
only transcribes rRNA genes
makes ribosomes
RNA polymerase 2
transcribes genes into mRNA
RNA polymerase 3
only transcribes tRNA genes
each has a specific promoter sequence it recognizes

12. Eukaryotic transcription
Which gene is read?
Promoter region
binding site before beginning of gene
TATA box binding site
binding site for RNA polymerase & transcription factors
Enhancer region
binding site far upstream of gene
turns transcription on HIGH

13. Transcription Factors
Eukaryotic transcription
Transcription Factors
Initiation complex
transcription factors bind to promoter region
suite of proteins which bind to DNA
hormones?
turn on or off transcription
triggers the binding of RNA polymerase to DNA

14. Matching bases of DNA & RNA
Match RNA bases to DNA bases on one of the DNA strands C U G A G U G U C U G C A A C U A A G C
RNA
polymerase U 5' A 3' G A C C T G G T A C A G C T A G T C A T C G T A C C G T

15. Post-transcriptional processing
mRNA’s require EDITING before use
primary transcript = pre-mRNA
mRNA splicing
INTRONS are removed (in between)
EXONS stay in message (expressed)
make mature mRNA transcript
Image by Riedell

16. Starting to get hard to define a gene!
Alternative splicing
Alternative mRNAs produced from same gene
when is an intron not an intron…
different segments treated as exons
Starting to get hard to define a gene!

17. EX: antibody production
Immune system needs to be able to make a huge number of different antibodies to match new and different invaders
Allows cell to use “old gene” in “new ways”
Able to respond to changing environment

18. mRNA EDITING snRNPs Spliceosome PROCESSING RNA SPLICEOSOMES
small nuclear RNA’s
proteins
Spliceosome
several snRNPs
recognize splice site sequence
cut & paste gene
THIS BREAKS THE RULES!
ALL ENZYMES ARE PROTEINS????
RIBOZYMES-RNA molecules
that function as enzymes (In some organisms pre-RNA
can remove its own introns)

19. More post-transcriptional processing
Need to protect mRNA on its trip from nucleus to cytoplasm
enzymes in cytoplasm attack mRNA
protect the ends of the molecule
add 5 GTP cap
add poly-A tail
longer tail, mRNA lasts longer: produces more protein
eukaryotic RNA is about 10% of eukaryotic gene. A
3' poly-A tail
mRNA 5'
5' cap 3' G P
A’s

20. Figure 12–18 Translation
Section 12-3

21. from nucleic acid language to amino acid language
Translation
from nucleic acid language to amino acid language

22. The code Code for ALL life! Code is redundant Start codon Stop codons
strongest support for a common origin for all life
Code is redundant
several codons for each amino acid
3rd base “wobble”
Why is the wobble good?
Strong evidence for a single origin in evolutionary theory.
Start codon
AUG
methionine
Stop codons
UGA, UAA, UAG

23. Transfer RNA structure
“Clover leaf” structure
anticodon on “clover leaf” end
amino acid attached on 3 end

24. tryptophan attached to tRNATrp tRNATrp binds to UGG condon of mRNA
Loading tRNA
Aminoacyl tRNA synthetase
enzyme which bonds amino acid to tRNA
bond requires energy
ATP  AMP
bond is unstable
so it can release amino acid at ribosome easily
The tRNA-amino acid bond is unstable. This makes it easy for the tRNA to later give up the amino acid to a growing polypeptide chain in a ribosome.
Trp
C=O
Trp
Trp
C=O OH
H2O OH O
C=O O
activating
enzyme
tRNATrp A C C U G G
mRNA
anticodon
tryptophan attached to tRNATrp
tRNATrp binds to UGG condon of mRNA

25. Protein synthesis/quiz
Ribosomes
Facilitate coupling of tRNA anticodon to mRNA codon
organelle or enzyme?
Structure
ribosomal RNA (rRNA) & proteins
2 subunits
large
small E P A

26. Ribosomes A site (aminoacyl-tRNA site) P site (peptidyl-tRNA site)
translation
Ribosomes
A site (aminoacyl-tRNA site)
holds tRNA carrying next amino acid to be added to chain
P site (peptidyl-tRNA site)
holds tRNA carrying growing polypeptide chain
E site (exit site)
empty tRNA leaves ribosome
goes to get recharged
Met U A C 5' A U G 3' E P A

27. Building a polypeptide1 2 3
How translation works
Building a polypeptide
Initiation
brings together mRNA, ribosome subunits, initiator tRNA
Elongation
adding amino acids based on codon sequence
Termination
end codon
Leu
Val
release
factor
Ser
Met
Met
Met
Met
Leu
Leu
Leu
Ala
Trp
tRNA C A G C U A C 5' U A C G A C U A C G A C G A C 5' A 5' U A A U G C U G A U A U G C U G A A U A U G C U G A A U 5' A A U
mRNA A U G C U G 3' 3' 3' 3' A C C U G G U A A E P A 3'

28. start of a secretory pathway
Protein targeting
Destinations:
secretion
nucleus
mitochondria
chloroplasts
cell membrane
cytoplasm
etc…
Signal peptide
address label
start of a secretory pathway

29. POST-TRANSLATIONAL MODIFICATIONS
Some amino acids modified by addition of sugars, lipids, phosphate groups, etc
Enzymes can modify ends, cleave into pieces
join polypeptide strands (4’ structure)
Ex: Made as proinsulin
then cut
Final insulin hormone
made of two chains
connected by
disulfide bridges

30. Post Translational regulation
UBIQUITIN - “death tags” for proteins
PROTEASOMES- recognize tags and destroy proteins

31. Can you tell the story? RNA polymerase DNA amino acids tRNA pre-mRNA
exon
intron
tRNA
pre-mRNA
5' GTP cap
mature mRNA
aminoacyl tRNA synthetase
poly-A tail 3'
large ribosomal subunit
polypeptide 5'
tRNA
small ribosomal subunit E P A
ribosome

32. Protein Synthesis in Prokaryotes
Bacterial chromosome
Protein Synthesis in Prokaryotes
Transcription
mRNA
Psssst… no nucleus!
Cell
membrane
Cell wall

33. Prokaryote vs. Eukaryote genes
Prokaryotes
DNA in cytoplasm
circular chromosome
naked DNA
no introns
Eukaryotes
DNA in nucleus
linear chromosomes
DNA wound on histone proteins
introns vs. exons
Walter Gilbert hypothesis:
Maybe exons are functional units and introns make it easier for them to recombine, so as to produce new proteins with new properties through new combinations of domains.
Introns give a large area for cutting genes and joining together the pieces without damaging the coding region of the gene…. patching genes together does not have to be so precise.
introns come out!
intron = noncoding (inbetween) sequence
eukaryotic
DNA
exon = coding (expressed) sequence

34. Translation in Prokaryotes
Transcription & translation are simultaneous in bacteria
DNA is in cytoplasm
no mRNA editing
ribosomes read mRNA as it is being transcribed

35. Translation: prokaryotes vs. eukaryotes
SEE PROCESSING VIDEO
Translation: prokaryotes vs. eukaryotes
Differences between prokaryotes & eukaryotes
time & physical separation between processes
takes eukaryote ~1 hour from DNA to protein
no RNA processing

36. COMPLETING PROTEINS POLYRIBOSOMES (POLYSOMES)
Numerous ribosomes translate same mRNA at same time
3-D folding (1’, 2’, 3’ structure)
Chaparonins help fold into 3-D shape