DNA makes ______ makes __________.

1
CHAPTER 17 FROM GENE TO PROTEIN

• DNA makes ______ makes __________.
• Proteins are the links between ________
  and_________.

• In 1909- Archibald _______ suggested that
• 1. genes dictate phenotype through enzymes that
  catalyze specific chemical reactions in the cell.
• 2. the symptoms of an inherited disease reflect a
  persons inability to synthesize a particular
  ___________.
• Gerrod speculated that alkaptonuria, a hereditary
  disease, was caused by the absence of an enzyme
  in a pathway

2
1. The study of metabolic defects provided
evidence that genes specify proteins

• The idea of __________ pathways was suggested
• 1930s- George Beadle and Boris Ephrussi
  speculated that each mutation affecting ________
  in Drosophila blocks pigment synthesis at a
  specific step by preventing production of the
  ___________ that catalyzes that step.

Actual evidence for metabolic pathways came from
Beadle and Tatum in the 1930s
3
Beadle and Tatum Experiment

• Organism used- -bread mold, Neurospora crassa.

Experiment
1. Mutate Neurospora with __________ 2. Screened
the survivors for mutants that differed in their
nutritional needs.

• ____________ Neurospora can grow on a
  ____________ medium
• Minimal medium agar, inorganic salts, glucose,
  and the
• vitamin biotin.
• Most nutritional mutants can survive on a
  complete growth medium which includes all 20
  amino acids.

3. If mutant failed to grow on minimal medium,
add amino acid ____________ until growth is
evident
Results
1. Identify a number of mutants that did not grow
on arginine, but would grow if supply arginine
pathways intermediates
4

• Their results provided strong evidence for the
  onegene - one __________ hypothesis.

Later modified to the one gene-one __________
hypothesis Why? Not all proteins are
_____________
Gene A
Gene B
Gene C
Gene A mutants
Gene B mutants
Gene C mutants
Fig. 17.2
5
2. Transcription and translation are the two main
processes linking gene to protein

• The bridge between DNA and __________synthesis is
  RNA.
• RNA differs from DNA
• 1. RNA contains ________ as its sugar (not
  deoxyribose)
• 2. _________ replaces thymine.
• AGTCAT becomes AGUCAU
• 3. An RNA molecule almost always consists of a
  ________ strand.

6

• In DNA or RNA, only four nucleotides produce
  life
• The specific _________ of hundreds or thousands
  of nucleotides in each gene carries the
  information for the primary structure of a
  protein, the linear order of the ____ possible
  amino acids.
• To get from DNA, written in one chemical
  language, to protein, written in another,
  requires two major stages, ____________
  and____________ .

7

• ____________ - DNA is the template for RNA,
  usually __________ RNA (mRNA).

• ____________________ - the information contained
  in the order of nucleotides in mRNA is used to
  determine the __________ sequence of a
  polypeptide.
• -Translation occurs at ____________.

Fig. 17.3a

• The basic mechanics of transcription and
  translation are ________ in eukaryotes and
  prokaryotes.

8
Major differences between prokaryotes and
eukaryotes-

• Transcription and translation are __________in
  bacteria (which lack________ ), but occur at
  separate locations in eukaryotes.
• - In bacteria, ___________attach to the leading
  end of a mRNA molecule while transcription is
  still in progress.
• -In a eukaryotic cells, almost all transcription
  occurs in the nucleus and translation occurs
  mainly at ribosomes in the cytoplasm.

Protein
RNA
DNA

• 2. Eukaryotes- Before the ________ can leave the
  nucleus it is modified in various ways during
  __________________ before the finished mRNA is
  exported to the cytoplasm.

Fig. 17.3b
9
3. In the genetic code, nucleotide triplets
specify amino acids

• A single or doublet code can not provide enough
  ____________ (4 and 16 respectively) to code for
  all 20 amino acids.
• Triplets of nucleotide bases are the smallest
  units of uniform length that can code for all the
  amino acids.
• In the__________, three consecutive bases specify
  an amino acid, creating 43 (64)
  possible__________.
• The genetic instructions for a polypeptide chain
  are written in DNA as a series of three-_________
  words.

10

• During transcription, one DNA strand,
  the_____________, provides a template for
  ordering the sequence of nucleotides in an RNA
  transcript.
• The _____________RNA molecule is synthesized
  according to base-pairing rules, except that
  _______ is the complementary base to adenine.
• During translation, blocks of three nucleotides,
  _______, are decoded into a sequence of amino
  acids.

Template strand
mRNA
Protein
Fig. 17.4
11

• During translation
• _________ are read in the 5-gt3 direction
• Each codon specifies _____ of the 20 amino acids
• It is a triplet code three bases for one amino
  acid
• It would take at least ____ nucleotides to code
  for a polypeptide that is 100 amino acids long.

Problem We have 64 possible combinations of the
nucleotides
Why 64 possibilities? How many ways can you
arrange 4 bases in sets of three? Answer
_____ ________________
Thus, predict that _______________ combination
must specify a given amino acid
12

• 1960s-Marshall Nirenberg determined the first
  match, that UUU coded for the amino acid
  _______________.
• Experiment - Add poly-U (uracil-only) RNA _______
• plus amino acids, ribosomes, other components.
• Result- This poly(U translated into a long chain
  of phenyalanine.
• Other more elaborate techniques were required to
  decode mixed triplets such a AUA and CGA.
• By the mid-1960s the entire code was
  _________________.

PhePhePhePhe
UUUUUUUUUUUUUUU
13
The genetic code
Know how to read this chart!!

• 1. ___ of 64 triplets code for amino acids.
• AUG codes for the methionine and _______ of
  translation.

• Three codons UAA, UAG and ______ do not code
  amino acids but signal the termination of
  translation.

• 2. The genetic code is ___________ but
  not______________ .
• Typically several different codons specify a
  given amino acid
• Any one codon indicates ___________ amino acid.

Fig. 17.5

• If you know a specific codon, you know the amino
  acid
• If you know only the amino acid, there may be
  several possible codons
• Example- Both GAA and GAG specify glutamate, but
  no other amino acid.

14
The genetic code (cont.)
Fig. 17.5

• Codons synonymous for the same amino acid often
  differ only in the _______ codon position.

Example
GUU, GUC, GUA and GUG all encode ________
A ________________ is established at the
translation start
3
RNA
5
UUACGAUGGAUUCAAACGUCAGGGCCUAAGGCUAG
Asp
Met
Asn
Ser
Arg
Val
Ala
Stop codon
Start codon
Summary- The genetic code uses____________________
______, or codons, each of which is translated
into a specific amino acid.
15

• The genetic code is nearly__________ , from
  bacteria to mammals

Thus, we can synthesize bacterial proteins
in_________
Exceptions do exist- they use slightly altered
genetic codes 1.single-celled eukaryotes like
Paramecium.
2. certain mitochondria and chloroplast_______
16
4. Transcription is the DNA-directed synthesis of
RNA

• Transcriptioncan beseparatedinto
  threestages1. _________2. elongation 3.
  __________

Fig. 17.7
17
What actually makes the RNA?

• Messenger RNA is transcribed from the template
  strand of a gene by _____________________.
• ___________________
• separates the DNA strands
• bonds the RNA nucleotides as they base-pair along
  the DNA template.
• can add nucleotides ______ to the ________ of the
  growing polymer.
• Genes are read _______ creating a ______RNA
  molecule.

RNA
5
3
DNA
3
5
18
Compare prok and euk polymerases

• Bacteria -________ type of RNA polymerase that
  synthesizes all RNA molecules.
• Eukaryotes -_________ RNA polymerases (I, II,
  and III) in their _________ .
• ___________________ is used for ________synthesis.

What marks the start of transcription??

• Answer- Specific sequences of nucleotides called
  the _________ mark where gene transcription begins

• In prokaryotes, RNA polymerase can recognize and
  bind __________ to the promotor region.

• In eukaryotes, proteins called ________________
  first bind the promotor region, especially
  a_________ , then RNA polymerase II binds

19
1. Initiation

• Eukaryotes- The complex including RNA polymerase
  II plus transcription factors is called the
• _________________.

Fig. 17.8
20
2. Elongation

• As RNA polymerase ________ the double helix,10 to
  20 bases at time.
• The enzyme addsnucleotides to the___ end of
  thegrowing strand.
• Behind the pointof RNA synthesis,the double
  helix_______ and theRNA moleculepeels away.

Fig. 17.7
21
2. Elongation (cont.)

• A single gene can be transcribed simultaneously
  by _____________ RNA polymerases at a time.
• A growing strand of RNA trails off from
  each______________.

RNA
5
5
DNA
3
TATAA
5
22
3. Termination

• Prokaryotes- RNA polymerase stops transcription
  at the end of the______________ .
• Both the RNA and DNA is then released.
• ______________- the polymerase continues for
  hundreds of nucleotides past the terminator
  sequence,_____________ .
• At a point about _____________ nucleotides past
  this sequence, the pre-mRNA is cut.

5
5
3
AAUAAA
TATAA
10-35 nucleotides
23
5. Eukaryotic cells modify RNA after transcription

• Modifications include
• 1. A ________at the 5 end of the pre-mRNA
  molecule
• The cap is a modified form of guanine
• Function a. Protect mRNA from
  __________enzymes.
• b. Attach here signal for____________

2. _____________50 to 250 adenine nucleotides at
the 3 end Function- a. inhibiting hydrolysis,
b. facilitating ribosome ___________c. facilitate
the export of mRNA from the nucleus.
3 tail
5 cap
Fig. 17.9

• The mRNA molecule also includes
  ____________leader and trailer segments.

24
RNA modification in eukaryotes (cont.)

• 3. ______________ - Most eukaryotic genes and
  their RNA transcripts have long ____________
  stretches of nucleotides.
• The noncoding segments are called ________
• The coding regions (final mRNA transcript) are
  called__________

• What is a coding region??
• RNA sequences that are translated into amino
  acid sequences

Coding region
Leader
Trailer
3 tail
5 cap
Fig. 17.9
25
Fig. 17.10

• RNA splicing removes ______ and joins ______
  to create an mRNA molecule with a _____________
  coding sequence.

26
This splicing is accomplished by
a________________.

• Contains small nuclear ribonucleoproteins
  (snRNPs). and small nuclear RNA molecules
  (snRNA). Each is about 150 nucleotides long.

Fig. 17.11
Splicing steps
(1) Pre-mRNA combines with _________ and other
proteins to form a spliceosome. (2) Within the
spliceosome, ________ base-pairs with nucleotides
at the ends of the intron. (3) The RNA
transcript is cut to release the________ , and
the exons are spliced together
Here the snRNA acts as a____________ , an RNA
molecule that functions as an enzyme.
27
Poly-A tail
Cap
28
6. Translation is the RNA-directed synthesis of a
polypeptide
How do proteins read the RNA molecule??
Answer-the tRNA molecule

• ______________(tRNA) - transfers amino acids
  from the cytoplasms pool to a ______________ .
• The ribosome adds each amino acid carried by
  tRNA to the growing end of the ______________
  chain.

Fig. 17.13
29

• A tRNA molecule
• Is about ___ nucleotides long
• Contains attachment site at the 3 end for an
  amino acid.
• Contains a loop with the ___________

Fig. 17.14
5
3
The anticodon base-pairs with a complementary
codon on mRNA.
If the codon on mRNA is UUU, a tRNA with a
______ anticodon and a tRNA carrying
phenylalanine will bind to it.

• The anticodons of some tRNAs recognize more than
  one________.
• Why? Because the rules for base pairing between
  the third base of the codon and anticodon are
  ________ (called______________).

30
How do we explain this wobble??

• If each anticodon had to be a perfect match to
  each codon, we would expect to find ___ types of
  tRNA, but the actual number is about____ .
• At the wobble position, U on the _____________can
  bind with A or G in the third position of a
  codon.
• Some tRNA anticodons include a modified form of
  adenine, inosine, which can hydrogen bond with U,
  C, or A on the codon.

31
Wobble base pairing
Fig. 17.5
Leu
Wobble
AAU
AAU
UGGCGAUGUUAGUAUUGCAUGAGUUAGGUGACCAAGAU
Start
Leu
Leu
32
How the is the tRNA linked to the amino acid??

• Each amino acid is joined to the correct tRNA by
  ____________________________The 20 different
  synthetases match the 20 different________________
  .
• The synthetase catalyzes a covalent bond between
  them, forming _______________ or activated amino
  acid.

Fig. 17.15

• ______________ (protein rRNA) facilitate the
  specific coupling of the tRNA anticodons with
  mRNA codons.

phetRNA
Protein
Ribosomal subunits
1. Large
RNA
2. Small
33

• Each ribosome has a binding site for mRNA and
  three binding sites for tRNA molecules.
• The ________ holds the tRNA carrying the growing
  polypeptide chain.
• The ______carries the tRNA with the
  next__________.
• Discharged tRNAs leave the ribosome at
  the_________.

Fig. 17.16
34

• Translation can be divided into three _________
  1. initiation 2.
  ____________ 3. termination
• Both initiation and chain elongation require
  energy provided by the hydrolysis of________.

Translation

• 1. Initiation
• Small ribosomal subunit binds mRNA
• Initiator tRNA (with methionine) is attached to
  start codon
• Initiation factors bring in the large subunit
  such that the initiator tRNA occupies the P site.

Fig. 17.17
35

• 2. Elongation - ________ steps per amino acid
  added
• a. Codon recognition- an elongation factor
  assists hydrogen bonding between the mRNA codon
  under the A site with the corresponding anticodon
  of tRNA carrying the appropriate amino acid.
• This step requires the ____________ of two GTP.

Translation
b. _____________________- a ________ molecule
catalyzes the formation of a peptide bond between
the polypeptide in the P site with the new amino
acid in the A site
c. ____________ - the ribosome moves the tRNA
with the attached polypeptide from the A site to
the P site.
Note mRNA is read 5 -gt 3, codon by codon.
Fig. 17.18
36
Translation

• 3. Termination occurs when one of the three
  ______ codons reaches the ___ site.
• A ____________ binds to the stop codon and
  hydrolyzes the bond between the polypeptide and
  its tRNA in the P site.

Fig. 17.19
Other translation facts

• Multiple ribosomes, polyribosomes, may trail
  along the same mRNA.
• A ribosome requires less than a minute to
  translate an average-sized mRNA into a
  polypeptide.

Fig. 17.20
37
Post-translational modifications-

• Additions of________ , lipids, or phosphate
  groups to amino acids.
• Enzymes may remove some amino acids or cleave
  whole polypeptide chains.
• Two or more ______________ may join to form a
  protein.

38
7. Signal peptides target some eukaryotic
polypeptides to specific destinations in the cell

• Recall that some ribosomes reside in two
  locations-
• Free ribosomes are suspended in the ________ and
  synthesize proteins that reside in the__________
  .
• __________ ribosomes are attached to the
  cytosolic side of the_______________________.
• They synthesize proteins of the endomembrane
  system as well as proteins secreted from the cell.

39

• Translation in all ribosomes begins in the
  cytosol
• A polypeptide destined for the ________________
  system or for export has a specific
  ________________ (approx 20 amino acids) region
  at or near the leading end.
• A _________________________(SRP) binds to the
  signal peptide and attaches it and its ribosome
  to a receptor protein in the ER membrane.

The SRP leaves and protein synthesis resumes with
the growing polypeptide snaking across the
membrane into the cisternal space
Fig. 17.21
40
8. RNA plays multiple roles in the cell a review

• RNA is versatile
• ________ - carries info for protein production
• _______-transport amino acids in translation
• _______- part of ribosome, has role in
  translation
• ________ - splicing mRNA

41
9. Comparing protein synthesis in prokaryotes and
eukaryotes a review
Euks
Proks
DNA polymerase
Requires
Transcription and translation coupled?
RNA processing?
Protein targeting?
42
10. Point mutations can affect protein structure
and function

• __________ are changes in the genetic material of
  a cell (or virus).
• include large-scale mutations in which ______
  segments of DNA are affected (translocations,
  duplications, and inversions).
• A chemical change in just one base pair of a gene
  causes a_______________

Fig. 17.23
In sickle cell, a single T to A mutation changes
amino acid from glu to val

• ______________________ - alterations of
  nucleotides still indicate the same amino acids
  because of redundancy in the genetic code.
• Many other mutations cause no effect in function

43

• Other base-pair substitutions cause a readily
  detectable change in a protein.
• ____________mutations are those that still code
  for an amino acid but change the indicated amino
  acid.
• ____________mutations change an amino acid codon
  into a _____ codon, nearly always leading to a
  nonfunctional protein.

Fig. 17.24
44

• Insertions and ____________ are additions or
  losses of nucleotide pairs in a gene.
• These have a __________ effect on the resulting
  protein more often than substitutions do.
• Unless these mutations occur in multiples of
  three, they cause a ___________ mutation.
• All the nucleotides downstream of the deletion or
  insertion will be improperly grouped into codons.
• The result will be extensive missense, ending
  sooner or later in nonsense - premature
  termination.

Fig. 17.24
45

• ______________ are chemical or physical agents
  that interact with DNA to cause mutations.
• ___________ agents include high-energy radiation
  like X-rays and ultraviolet light.
• _________ mutagens may operate in several ways.
• As base ________ ___that may be substituted into
  DNA, but that pair incorrectly during DNA
  replication.
• Interfere with DNA replication by inserting into
  DNA and distorting the____________________.
• Cause chemical changes in bases that change their
  pairing properties.

46
11. What is a gene? revisiting the question

• ___________ concept - a discrete unit of
  inheritance that affects phenotype.
• Morgan and his colleagues assigned genes to
  specific loci on chromosomes.
• A specific nucleotide sequence along a region of
  a DNA molecule.
• A DNA sequence that codes for a specific
  polypeptide chain.
• A region of DNA whose final product is either a
  polypeptide or an RNA molecule.

or
or
or
47
Fig. 17.26