435
MCB 3020, Spring 2005
Chapter 7:
Molecular Genetics
Molecular Genetics I: Replication
I. Heredity and Genetics
II. Genomes
III. DNA structure
IV. Bacterial DNA replication
V. Replication at the ends of
linear DNA
436
I. Heredity
The transmission of characters
to progeny.
DNA carries the information
necessary for the transmission
of characters.
The biological information is encoded
in the sequence of bases.
TB
437
Genetics:
438
the study of the mechanisms
of heredity and variation in
organisms
TB
439
Flow of information
replication
DNA  DNA
transcription

RNA
translation

protein
II. Genome
Genome = all the DNA of a cell
(or all the genetic
material of a virus)
440
Typical bacterial genome
441
one circular doubleoften
stranded DNA chromosome plasmid(s)
500-12,000 genes
TB
typical viral genome
442
DNA
or
RNA
4-200 genes
TB
Typical eukaryotic genome
443
4-224, linear chromosomes
5,000 - 125,000 genes
TB
III. DNA structure
deoxyneucleotides
phosphodiester bonds
5' and 3' ends
antiparallel
complementary
double helix
444
TB
NH2
N
HOCH2
N
445
N
N
HO H
Deoxyadenosine (purine)
TB
H 3C
HOCH2
O
446
NH
N
O
HO H
deoxythymidine (pyrimidine)
TB
phosphodiester bond
447
-P-O-C
O
O-
O
P
O
C
ssDNA
-O
TB
448
ring numbering
system for
deoxyribose
5’
-C
1’
4’
5' end
-P-O-C
O
O-
3’ end
O
P
O
3’
2’
C
HO
ssDNA
TB
449
dsDNA
antiparallel
5’
3’
3’
5’
dsDNA is always antiparallel
TB
complementary
450
5’- GGATGCGT -3’
3’-CCTACGCA-5’
Two ssDNA molecules joined by
standard base-pairing rules
In dsDNA, the strands are always
complementary.
TB
451
double helix
right handed
TB
Supercoiling
relaxed DNA
452
supercoiled DNA
Within cells DNA is supercoiled
TB
IV. Bacterial DNA replication
453
DNA synthesis using a DNA template
Complementary base pairing
(A=T, GC) determines the sequence
of the newly synthesized strand.
DNA replication always proceeds from
5’ to 3’ end.
TB
454
Flow of information
replication
DNA  DNA
transcription

RNA
translation

protein
455
Overview of bacterial DNA replication
single origin (in bacteria)
bidirectional
theta structures
replication fork
semi-conservative
TB
456
bacterial DNA replication
bidirectional
origin (start point)
bacterial
chromosome
TB
457
two replication forks
theta
structure
TB
458
semi-conservative
*
*
*
+
*
TB
IV. Bacterial DNA replication
Key Enzymes
helicase
ssDNA binding protein
primase
DNA polymerase III
DNA polymerase I
DNA ligase
459
TB
Important facts
460
All DNA polymerases require a primer
DNA is synthesized 5' to 3'
TB
helicase
Unwinds duplex DNA
461
TB
ssDNA binding protein
462
binds to and stabilizes ssDNA
prevents base pairing
ssDNA binding protein
TB
463
primase
synthesizes a short RNA primer
using a DNA template
primase
RNA primer
(a short starting sequence
made of RNA)
TB
DNA polymerase III
464
Synthesizes DNA from a DNA
template and proofreads
TB
DNA polymerase I
465
Synthesizes DNA from a DNA
template and removes RNA primers.
TB
DNA ligase
Joins DNA strands together by
forming phosphodiester bonds
466
DNA ligase
TB
replication fork
5'
467
lagging strand
3'
5'
leading strand
template strands
3'
TB
Leading strand
synthesis
5'
468
RNA primer
helicase
ssDNA binding proteins
3'
TB
5'
469
DNA polymerase
helicase
ssDNA binding proteins
3'
TB
Leading strand synthesis
5'
470
DNA pol III
helicase
DNA
ssDNA binding proteins
3'
TB
Lagging strand synthesis
(discontinuous)
471
Okazaki
fragment
3'
5'
(~1000 bases)
(primase)
helicase
ssDNA binding proteins
pol III
3'
TB
Primer removal
pol
III
3'
472
5'
pol I
pol I
5’ to 3’
exonuclease
activity
TB
473
Ligation
DNA ligase
TB
Proofreading
474
Pol III removes misincorporated bases
using 3' to 5' exonuclease activity
This decreases the error rate to about
-10
10 per base pair inserted
TB
475
V. Replication of the ends of linear DNA
Since all known DNA polymerases
need a primer, how are the ends of
linear DNA replicated in eukaryotes?
newly synthesized DNA
5'
template
RNA primer
3'
TB
476
Telomeres
repetitive DNA at the end of linear
eukaryotic chromosomes
Example
(GGGGTT)n
n = 20 - 200
GGGGTT GGGGTT GGGGTT
5'
TB
Telomerases are enzymes that add
DNA repeats to the 3' end of DNA.
Telomerases are
composed of protein and
an RNA molecule that
functions as the template
for telomere synthesis.
477
AACCCCAAC
telomerase
478
5'
AACCCCAAC
5'
GGGGTTGGGGTT
telomerase
479
AACCCCAAC
5'
GGGGTT GGGGTT GGGGTT
primase
5'
GGGGTT GGGGTT GGGGTT
pol III
5'
pol I
ligase
telomeric repeats
480
For most cells, telomeres are added
during development. Later
telomerase becomes inactive.
481
Hence, as cells divide the DNA
becomes shorter.
Note that telomerase is reactivated in
many types of cancer cells.
TB
Study objectives
482
1. Compare and contrast bacterial, viral and eukaryotic genomes.
2. What are the 4 bases in DNA? Which are purines? Which are pyrimidines?
What is the sugar? I will not ask you to recognize the structures of individual
bases, but note that deoxythymidine has a methyl group in the pyrimidine ring.
3. Understand how the following terms apply to DNA structure:
phosphodiester bonds, 5' and 3' ends, antiparallel, complementary,
double helix. What parts of the nucleotides are joined in the phosphodiester
bond?
4. Understand how the following terms apply to DNA replication:
template, complementary base pairing, origin, bi-directional, theta structures,
replication fork, semi-conservative.
5. Know how the following enzymes function in leading and lagging strand
replication: helicase, ssDNA binding protein, primase, DNA polymerase III,
DNA polymerase I. What is an Okazaki fragment?
6. What is proofreading?
7. Understand the problem of replicating the ends of linear DNA. Understand
how telomerase solves that problem for eukaryotic chromosomes.
Molecular Genetics II: Transcription
I. RNA
II. Gene expression
III. Prokaryotic Transcription
483
484
Flow of information
replication
DNA  DNA
transcription

RNA
translation

protein
I. RNA (ribonucleic acid)
485
A polymer of nucleosides held
together by phosphodiester bonds.
RNA is usually single-stranded.
RNA plays a key role in decoding
the information in DNA.
TB
A. Functions of the major RNAs
486
1. messenger RNAs (mRNA) contain
genetic information to encode a protein
phe
2. transfer RNAs (tRNA) act as adapters
between the mRNA nucleotide code and
amino acids during protein synthesis
3. ribosomal RNAs (rRNA) are structural and
catalytic component of ribosomes
B. RNA structure
487
1. RNA nucleosides
2. phosphodiester bonds
3. 5' and 3' ends
4. complementary base pairing
5. stem-loops
TB
1. The RNA nucleosides
488
The RNA nucleosides have
2'-hydroxyl groups which are
not found in DNA.
"U" is found in RNA (in place of "T")
Guanosine (G)
Adenosine (A)
Cytidine (C)
Uridine (U)
TB
489
2. The phosphodiester bonds
of RNA are analogous to those
of DNA.
3. The 5' and 3' ends
of RNA are analogous to
those of DNA.
TB
P-O-C
5' end
phosphodiester
bond
O
O-
490
ring numbering
system for ribose
5'
-C O
1’
4’
O
OH
O
P
3’ end
3’
O
C
HO
2’
O
OH
RNA
TB
4. Complementary base pairing
491
CCCTTTGGGAAA
DNA
GGGAAACCCUUU
RNA
GGGAAACCCUUU
RNA
CCCUUUGGGAAA
RNA
hydrogen
bonding
TB
5. RNA stem loops
492
A common RNA secondary structure
ssRNA
complementary
base pairing
(helical)
TB
493
II. Gene expression
Different scientists define the term
gene expression differently. Most
commonly, gene expression refers to
the decoding of genes into proteins
or RNAs.
1 gene encodes 1 polypeptide,
1 tRNA, 1 rRNA, or 1other RNA
TB
A. Gene numbers
494
group
approximate
gene number
viruses
prokaryotes
eukaryotes
4-200
500-12,000
5,000-125,000
TB
Any given species has a unique set
of genes that confers a unique set
of properties.
495
Proteins and RNAs determine all of the
characteristics of organisms and cells.
Example: Escherichia coli has 4405 genes
~117 encode RNAs (tRNA, rRNA)
~4288 encode proteins
TB
C. Gene expression in prokaryotes
1. Expression of single genes
496
Ex.1: a single gene that encodes a protein
1 gene
transcription
translation
1 mRNA
1 polypeptide
TB
Ex. 2: a single gene that encodes one
rRNA or tRNA
497
1 gene
transcription
1 RNA
RNA processing
degraded
1 tRNA etc.
TB
2. Expression of operons
operon
two or more genes
transcribed together
498
A
B
C
DNA
transcription
polycistronic message
polycistronic
mRNA
a single RNA molecule that
represents more than one gene
TB
a. Operons can encode several
polypeptides or proteins.
A
B
499
1 operon
C
transcription
1 polycistronic mRNA
translation
B
A
C
2 or more polypeptides
TB
b. Operons can encode several
rRNA molecules.
500
1 operon
1 polycistronic
RNA
processing
rRNA
rRNA
degraded
2 or more rRNAsTB
3. Important points
501
The details of organization,
processing and degradation are
different for different RNAs.
Most prokaryotes use operons.
Operons are used to coordinate
gene expression and often contain
genes of related function.
TB
D. Eukaryotic gene expression
502
1. Expression of eukaryotic
rRNA and tRNA genes
The expression of rRNA and
tRNA is similar in eukaryotes
and prokaryotes.
TB
2. Eukaryotic protein expression
503
a. Typical eukaryotic genes have exons and introns.
E I
E
I
E
I
E
gene
E = exon = coding sequences
I = intron = intervening,
noncoding sequences
Eukaryotes do NOT have operons
TB
1 gene with exons and introns
E I
E
I
E
I
504
E
transcription
1 RNA representing exons and introns
(primary transcript)
TB
b. Primary transcripts
505
primary transcript
processing
1 mRNA
1 polypeptide
TB
c. Processing of primary transcripts
506
i. capping
ii. splicing
iii. tailing
TB
i. Capping
507
Addition of a 5' cap
CAP
Capping usually occurs before
transcription is finished.
TB
CH3
Typical 5' CAP
P
P
P
N
OCH2 N
O
O
508
N
N
NH2
HO OH
5' carbon
of RNA chain
7-methylguanosine
5' to 5' linkage
Know the name (methylguanosine cap, 5' cap),
but don't memorize structure.
TB
ii. Splicing
The removal of introns.
509
primary transcript
splicing
RNA without introns
TB
iii. Tailing
510
Addition of a poly-A tail
A1A2...A~200
TB
3. Notes on eukaryotic
511
RNA processing
The exact order of capping, tailing
and splicing varies for different genes.
Processing occurs in the nucleus
Poly-A tails are added by poly-A
polymerase, NOT during transcription.
TB
E. Comparison of eukaryotic and
prokaryotic gene expression.
512
Eukaryotic mRNAs are usually
spliced,capped and tailed.
Eukaryotes do NOT have operons.
tRNA and rRNA expression are
generally similar
Prokaryotic genes very very rarely
have introns
TB
III. Prokaryotic transcription
513
A. overview
B. transcribed regions
C. RNA polymerase
D. promoters
E. terminators
F. sigma factor
TB
514
Flow of information
replication
DNA  DNA
transcription

RNA
translation

protein
515
A. Overview of prokaryotic transcription
RNA synthesis from a DNA template
typical
gene dsDNA
RNA polymerase
primary transcript complementary to
one strand of the coding region
TB
B. Defined regions are transcribed
516
upstream transcribed downstream
region
region
region
gene dsDNA
transcription
promoter
start
site
(RNA polymerase
binding site)
termination
site
TB
C. RNA polymerase is the enzyme that 517
synthesizes RNA from a DNA template.
RNA polymerase
gene,or
operon
DNA template
complementary RNA
TB
518
+
+
completed
transcript
TB
519
D. Promoters
Sites on DNA where RNA
polymerase binds to start transcription
upstream transcribed downstream
region
region
region
gene dsDNA
promoter
transcription
start site
termination
site
TB
1. Typical bacterial 70 promoter
TTGACA
AACTGT
TATAAT
ATATTA
TTGACA =
-35 consensus
sequence
TATAAT =
-10 consensus
sequence
520
*also called Pribnow box;
~ 10 bases before start
site of transcription TB
A more common way to draw a promoter 521
3'
5'
TTGACA
-35
TTAACT
-10
Note:
The - 10 and -35 sequences can vary somewhat.
TB
E. Transcriptional terminators
522
DNA region that mediates the
termination of transcription.
termination
site
gene dsDNA
region where
terminators are
usually found
TB
1. Intrinsic terminator
523
DNA encoding an RNA that forms
a stem loop followed by a run of "U"s
that is used for transcriptional termination.
RNA
UUUU
3' end of RNA
TB
Intrinsic terminator function
524
The RNA stem loop
binds to RNA pol and
causes termination
Important fact: Intrinsic terminators must be
transcribed in order to function.
TB
525
2. Rho-dependent terminator
A DNA site where RNA polymerase
pauses and transcription
is terminated by Rho protein
TB
526
Rho protein binds RNA then moves
along RNA until it contacts RNA pol and
terminates transcription
Rho protein
RNA pol
pauses at
Rho
termination site
TB
F. The sigma factor cycle
527
Sigma factors ( ) are a subunit
of RNA polymerase.
Sigma factors are needed for promoter
binding, but after transcription starts they
dissociate.
TB
1. Subunit structure of bacterial
RNA polymerase
core enzyme
'
528
holoenzyme
'
The holoenzyme includes one
of several sigma factors.
TB
RNA pol holoenzyme (core + sigma)
sigma
factor
529
core enzyme
sigma factor
RNA (~10 nucleotides)
TB
termination
530
RNA
+
core enzyme
sigma
holoenzyme
TB
Upstream region of the lactose operon531
TAATGTGAGTTAGCTCACTCATTA
-35 region
GGCACCCCAGGCTTGACATTTATG
-10 region (Pribnow)
CTTCCGGCTCGTATGTTGTGTGGA
Transcription
start site
AATTGTGAGCGGATAACAATTTCA
Shine-dalgarno (RBS)
CACAGGAAAGAGCTATGACC...
Translation start site
Study objectives
532
You will need to know ALL the concepts and details in this lecture.
1. What are the three main types of RNA and what are their functions?
2. Understand how the following terms apply to RNA structure: phosphodiester
bonds, 5' and 3 ends, nucleosides, complementary base pairing, stem loops.
3. Compare and contrast DNA and RNA structure.
4. What is a gene? What is gene expression? *Understand transcription,
translation, and RNA processing in both prokaryotes and eukaryotes.
5. Define operons and polycistronic messages. How do they function in
prokaryotic gene expression?
6. *Compare and contrast the features of prokaryotic and eukaryotic gene
expression. Do eukaryotes have operons? What are exons, introns, primary
transcripts, capping, tailing, and splicing. What is the 5' cap (methylguanosine
cap)? How and when is the poly-A tail added to the transcript? Where does
eukaryotic RNA processing occur?
7. Understand the structure and function of promoters and terminators in
transcription. Contrast intrinsic terminators and rho-dependent terminators.
8. Know the subunit structure of bacterial RNA polymerase and the sigma cycle.
Molecular Genetics III:
Prokaryotic translation
533
I. Key components of translation
II. Steps in translation
III. The genetic code
Overview of prokaryotic translation
534
Protein synthesis from an mRNA template.
translated region
mRNA
phe
translation
protein of specific amino acid sequence
TB
I. Key components of translation
A. mRNA
B. tRNA
C. ribosomes and rRNA
535
536
A. mRNA
RNA template for protein synthesis
translated
region
Shineseries
of
codons
Dalgarno
(usually
~300
codons)
sequence
mRNA
start codon
stop codon
TB
1. Shine-Dalgarno sequence
537
~AGGAGG, ribosome binding sequence,
critical for ribosome binding
2. start codons
AUG, GUG, or UUG
3. stop codons (nonsense codons)
UAA, UGA, or UAG
TB
538
4. Translated region (coding sequence)
• Series of codons that determines
the amino acid sequence of the
encoded protein.
• Coding sequences have an average
of about 300 codons.
• Except for the stop codon, each
codon specifies a particular amino
acid.
TB
5. Codons consist of 3 bases
539
start
codon
codons
protein
2
3 4
1
AUGCAUUGUUCU...
fMet - His - Cys - Ser ...
1
2
3
4 TB
B. tRNA
The adapter molecule for translation
540
1. Particular tRNAs carry
particular amino acids.
f-Met
tRNA-f-Met
His
His
tRNA-His
TB
2. Particular tRNAs recognize
particular codons.
codons
541
AUGCAUUGUUCU...
tRNAs
AA1
AA2
amino acid (AA)
This allows amino acids to be brought
TB
together in a particular order.
3. tRNA structure
All tRNAs are generally similar
in structure.
542
a. 1o structure
ssRNA 73-93 nucleotides long
5'
UAC
3'
TB
o
2
b. structure
clover leaf
543
acceptor arm
TYC arm
D-arm
extra arm
anticodon loop
TB
c.
o
3
structure
inverted "L"
544
TB
d. Anticodon
545
A 3 base sequence in tRNA
complementary to a specific codon.
anticodon
Base pairing between an anticodon and a codon
allows a tRNA to recognize a specific codon. TB
e. codon-anticodon interactions
anticodon
3'
5'
321
5'
546
UUA
AAU
123
tRNA
3' mRNA
codon
TB
4. tRNA charging (adding amino acid)547
O
H2N-CH-C-O 3'
R
3'
tRNA
(uncharged)
aminoacyl-tRNA
(charged)
tRNA charging uses the energy of ATP TB
Aminoacyl-tRNA synthetases
548
enzymes that attach amino acids to tRNA
enzyme
ATP
amino acid
aminoacyl-AMP
tRNA
aminoacyl-tRNA
PPi
AMP
TB
AMP = adenosine monophosphate
PPi = inorganic pyrophosphate
5. tRNA facts
549
Prokaryotes have about 60 different
tRNAs.
tRNAs contain many modified bases.
TB
C. Ribosomes and rRNA
550
Ribosomes
ribonucleoprotein complexes that
catalyze protein synthesis.
rRNAs
have structural and catalytic roles
TB
1. Prokaryotic 70s ribosome
551
23s rRNA
5s rRNA
34 proteins
50s
subunit
16s RNA
21 proteins
30s
subunit
TB
2. Ribosomal sites where tRNAs bind
552
E = exit
E
P
A
P = peptidyl
A = aminoacyl
TB
3. 16S rRNA
The 3' end of the 16s rRNA is
complementary to the ShineDalgarno sequence (ribosome
binding sequence of mRNAs)
553
II. Steps in translation
A. initiation
P-site
30s subunit
of ribosome
554
f-met
AGGAGG-----AUG
Shine-Dalgarno
(AGGAGG
on mRNA)
f-met
mRNA
AUG
GTP hydrolysis
50s subunit
30s subunit TB
1. f-met tRNA (formyl-methionine tRNA)
555
In Bacteria, different met-tRNAs are used for
elongation and initiation.
met
tRNA
initiation, formyl-methionine
met
tRNA
elongation, methionine
f
m
TB
2. Initiation in different domains
556
In Bacteria, the formyl group of the initiator
formylmethionine (f-met) is later removed.
In Eukarya and Archaea, initiation begins
with methionine rather than f-met.
In Eukarya, the ribosome recognizes
the 7-methylguanosine cap at the 5’ end of
mRNA and initiates at the first AUG.
TB
B. Elongation
557
1. AA-tRNA binding
AA
AA
mRNA
P-site
A-site
AA AA
TB
2. peptide bond synthesis
558
AA AA
(peptidyl transferase)
AA
AA
TB
3. translocation
AA
AA
GTP
hydrolysis
559
AA
AA
TB
C. Termination
560
AA AA AA AA
stop codon
AA
UAA
termination
AA AA AA AA
AA
TB
D. Additional notes on translation
561
1. Ribosomes move along the mRNA.
mRNAs can be translated by 5-10
ribosomes simultaneously.
mRNA
"Polysomes" are mRNAs with
several ribosomes attached.
TB
2. In prokaryotes only, transcription
and translation are coupled.
562
Translation begins before
transcription ends.
DNA
mRNA
TB
563
3. Protein folding into the active form
can occur spontaneously or with the
help of a large protein complex
called a molecular chaperone.
ATP
ADP
improperly
folded protein
molecular
chaperone
properly folded protein
III. The genetic code
A. universal code
B. degenerate code
1. synonyms
2. codon families
3. codon pairs
C. wobble base pairing
564
III. The genetic code
565
8 codon families, 14 codon pairs, 3 stop codons
(Do not
memorize)
A. The genetic code is almost
universal.
566
Most organisms use the same
genetic code.
TB
B. The genetic code is degenerate.
567
more than one codon can code
for the same amino acid
UUU  phenylalanine
UUC  phenylalanine
TB
1. synonyms
codons that code for the same
amino acid
UUU  phenylalanine
UUC  phenylalanine
Not all synonyms are used with
equal frequency. This is called
"codon usage bias".
568
569
2. codon families
CUU
CUC
CUA
CUG
any nucleotide in
the 3rd positions
leucine
TB
3. codon pairs
any pyrimidine in
the 3rd position
UUU
UUC
phenylalanine
CAA
CAG
glutamine
any purine in
the 3rd position
570
TB
C. Wobble base pairing
571
U-G and G-U base pairs are allowed in
the 3rd position of the codon.
codon (mRNA)
5'
3'
UUU
AAG
anticodon (tRNA)
3'
5'
TB
572
Flow of information
replication
DNA  DNA
transcription

RNA
translation

protein
Study objectives
573
1. Know the DETAILS of the structure and function of mRNA, tRNA, rRNA, and
ribosomes in translation. Memorize the start and stop codons. You do NOT
need to memorize codons other than the start and stop codons.
2. What reaction is catalyzed by aminoacyl-tRNA synthetases?
3. For the process of translation, know the details of initiation, elongation
peptide bond formation, translocation and termination.
4. Compare and contrast Bacterial, Archaeal and Eukaryal initiation.
5. What are polysomes?
6. What is meant when it is said that transcription and translation are coupled
in prokaryotes?
7. Some proteins fold spontaneously while others require assistance.
What are molecular chaperones?
8. How do the following terms apply to the genetic code: synonyms,
codon pairs, codon families, wobble, codon usage bias.
574
MCB 3020, Spring 2004
Chapter 7:
Regulation of
Gene Expression
Regulation of Gene Expression I:
I. Regulation of gene expression
II. Transcriptional regulation
III. Examples of gene repression
IV. Example of gene induction
575
I. Regulation of gene expression
576
Not all genes are turned on
(expressed) all the time
In general, they are turned on
only when needed.
TB
Cells can respond to environmental 577
changes by regulating gene expression.
arginine
maltose
lactose
glucose
tryptophan
Different genes are expressed when
cells grow on different compounds.
glucose
e.g. Growth on
lactose requires
expression of at
least three
additional genes.
578
maltose
TCA
lactose
(galactose--1,4-glucose)
P O
lacZ
-galactosidase
lacY
lacA
lac permease (transport protein)
A. Why regulate gene expression?
579
Regulation allows cells to respond
to environmental conditions by
synthesizing selected gene products
only when they are needed.
B. Gene expression
synthesis of a gene product
1. constitutive
2. regulated
580
1. Constitutive gene expression
expression of genes at about the
same level under all
environmental conditions
581
e.g. "housekeeping genes" like
primase
ssDNA binding proteins
TB
2. Regulated gene expression
Control of the rate of protein
or RNA synthesis as an adaptive
response to stimuli.
582
induction: increase in gene expression
repression: decrease in gene expression
a. gene induction
increase in gene expression
amount of
gene product
583
inducer
time
TB
584
e.g. genes that encode maltose-utilizing
enzymes are induced by maltose.
maltose absent
maltose added
maltose
catabolic
enzymes
(molecules/cell)
lag phase
time
b. gene repression
decrease in gene expression
585
amount of
gene product
time
TB
e.g. genes that encode enzymes for
tryptophan biosynthesis are
repressed by tryptophan.
tryptophan absent
tryptophan present
enzymes for
tryptophan
biosynthesis
(molecules/cell)
time
586
Important general principle
587
• catabolic substrates (e.g. maltose
and lactose) induce the genes
required for their catabolism
• biosynthetic molecules (e.g. amino
acids and purines) repress the
genes required for the biosynthesis
II. Transcriptional regulation
588
• regulation of RNA synthesis
• the most common method of
gene regulation in all cells
A. Regulatory proteins
B. Regulatory protein binding sites
C. Effector molecules
TB
A. Regulatory proteins
589
• Transcriptional regulation is
mediated by regulatory proteins.
• Cells have many different regulatory
proteins.
• Specific regulatory proteins control the
transcription of specific groups of genes.
• Examples of regulatory proteins are
"repressor proteins" and "activator proteins."
TB
1. Repressor proteins
590
Repressor protein (dimer)
DNA
P
RNA polymerase
Promoter
Repressor proteins decrease transcription
when bound to DNA by interfering with the
TB
activity or binding of RNA polymerase.
2. Activator proteins
Activator protein
P
591
DNA
RNA polymerase
"weak" promoter
Activator proteins increase transcription when
bound to DNA by helping RNA polymerase bind
TB
to weak promoters.
B. Regulatory protein binding sites
592
Regulatory proteins bind to specific
DNA sequences.
A particular regulatory protein will
only control the expression of
genes having appropriate binding
sites.
TB
1. Operator sites
593
binding sites for repressor proteins
GTGTAAACGATTCCAC
CACATTTGCTAAGGTG
lac
repressor
binding site
Imperfect palindrome
Usually found near promoters.
TB
2. Activator binding sites
Binding sites for activator proteins
GTGAGTTAGCTCAC
CACTCAATCGAGTG
594
crp
binding
site
Imperfect palindrome
Usually found near promoters.
TB
C. Effector molecules
595
Small molecules from the
environment (or made inside cells)
that signal specific changes
in gene expression.
TB
1. Classes of effectors
a. inducers
596
maltose
small molecules that mediate
gene induction
e.g. catabolic substrates:
sugars, amino acids, fatty acids
lactose
TB
b. corepressors
small molecules that mediate
gene repression
597
e.g. biosynthetic products:
amino acids, purines, pyrimidines,
fatty acids etc.
arginine
tryptophan
TB
2. How effectors work
598
Effectors change the DNA binding affinity
of regulatory proteins for their binding sites.
regulatory protein
effector
conformational change
(change in 3-D structure)
TB
A. Some effectors increase
DNA binding affinity
599
regulatory protein
conformational change
(change in 3-D structure)
DNA
effector
TB
B. Some effectors decrease
DNA binding affinity
600
regulatory protein
DNA
conformational change
(change in 3-D structure)
effector
TB
601
Since most regulatory proteins influence
transcription when bound to DNA,
the binding of effectors to regulatory
proteins changes gene expression.
regulatory
protein
effector
TB
III. Examples of gene repression
602
A. Regulation of the trp operon
B. Regulation of the arg operon
TB
A. The trp operon is a group of genes used for603
biosynthesis of the amino acid tryptophan (Trp).
The trp operon
trp genes
promoter
E
D
C
B
A
polycistronic mRNA
Five enzymes for tryptophan biosynthesis TB
604
1. When Trp is NOT available in the
environment, expression of the trp
operon allows Escherchia coli to
make Trp needed for protein
synthesis.
2. When Trp is available, E. coli
takes up Trp from the environment
and represses the trp operon.
TB
605
trp promoter
inactive
repressor
operator
tryptophan
active
repressor
RNA polymerase
genes on
genes off
TB
Note: Repression of the trp operon by606
tryptophan involves a repressor protein.
• When tryptophan binds to the repressor
protein, the repressor protein binds to DNA.
• Transcription is blocked.
Result: VERY low amounts of tryptophan
are synthesized when the cell can get
tryptophan from the environment .
B. Regulation of the arg operon
for arginine biosynthesis
607
If arginine is present in large amounts
• arg biosynthetic enzymes NOT needed
• arg binds repressor
• arg-repressor binds DNA
• RNA polymerase can't bind to promoter
argC
argB
argH
operator
arg biosynthetic genes
 transcription rate decreases
P
608
If arg is absent, the cell needs to make arg
• repressor doesn't bind DNA
• RNA polymerase can bind
• transcription of arg genes occurs
P
operator
argC
argB
argH
arg biosynthetic genes
IV. Example of gene induction:
Regulation of the lac operon
609
A. The lac operon is a group of genes
used for catabolism of the sugar lactose.
lac genes
promoter
Z
Y
A
operator
TB
• When lactose is unavailable, the
catabolic enzymes are NOT
needed.
The lac operon is
expressed at only very low levels.
• When lactose is available, E. coli
induces expression of lac operon.
610
TB
611
B. Lactose unavailable
lac promoter
Z
Y
A
genes
off
In the ABSENCE of lactose,
the lac repressor protein binds DNA.
Note: the role of crp/cAMP in control of the
lac operon is not considered here.
TB
C. Lactose available
612
lac promoter
Z
Y
A
genes
on
RNA polymerase
lactose
allolactose
repressor does not
bind DNA TB
Important points
613
Repressor proteins can mediate
gene repression (e.g. trp operon)
or gene induction (lac operon).
Activator proteins can mediate both
gene induction and gene repression.
TB
614
Some repressor proteins mediate gene induction.
Example: the lac repressor
Lactose (a sugar) can be an energy source.
If lactose is absent,
• enzymes for using lactose are not needed
• lac repressor binds to the lac operator
• the lac genes are not expressed
CAP
site
P O
lacZ
lacY
lacA
615
Some repressor proteins mediate gene induction.
Lactose ( ) induces the expression of lac genes.
If lactose is present,
+
• enzymes for using lactose are needed
• (allo)lactose binds to the lac repressor and
causes a conformational change
• repressor-lac does NOT bind to DNA
• expression of lac genes is possible
CAP
site
P O
lacZ
lacY
lacA
Study objectives:
Please study all the concepts and details of Regulation.
616
1. Why do cells control gene expression? What is constitutive gene expression?
2. What is gene induction? gene repression?
3. Are catabolic genes more likely to be repressed or induced? Why?
4. Are anabolic (biosynthetic) genes more likely to be repressed or induced?
Why?
5. What are the functions of the following in the regulation of transcription:
repressor protein, activator protein, effector, co-repressor, inducer,
activator binding site, operator, palindromic sequences,
protein conformational changes? Understand the concepts and details.
Do NOT memorize the specific palindromic sequences.
6. Describe regulation of the trp operon and arg operon by repressor proteins.
7. Describe the effect of lactose on the induction of the lac operon.
8. Explain how repressor proteins can mediate gene repression.
Explain how repressor proteins can mediate gene induction.
9. Know that some activator proteins can mediate gene induction,
while other activator proteins mediate gene repression
617
Regulation of Gene Expression II:
I. Activator proteins
II. Global regulation
III. Two-component regulatory systems
IV. Attenuation
I. Activator proteins
618
Proteins that activate
transcription when bound to
activator binding sites.
eg. maltose activator protein
catabolite activator protein (CAP)
cyclic AMP receptor protein (crp)
619
A. Typical activator protein
DNA
P
activator
binding site
RNA polymerase
unusual promoter
TB
B. Typical activator binding site
P
620
O
GTGAGTTAGCTCAC
CACTCAATCGAGTG
Imperfect palindrome
TB
C. Unusual promoters are involved
in control by activator proteins.
No
-35
consensus
621
-10
consensus
(Pribnow box)
TB
D. The catabolite activator protein
1. In the lac operon, the activator
protein is called the catabolite
activator protein (CAP) or the
cyclic AMP receptor protein (crp).
2. When cyclic AMP (cAMP) is
present, the cAMP/CAP (crp)
complex binds DNA and
activates transcription.
622
CAP
(crp)
cAMP
cAMP/CAP
complex
binds DNA TB
NH2
N
O
CH2
N
623
N
N
(Don't
memorize)
HOP=O
O
OH
cyclic AMP (cAMP)
cyclic adenosine monophosphate
TB
Role of CAP (crp) in the lac operon
624
Without activator protein, RNA polymerase
binds weakly and the transcription rate is low.
crp
P O
binding site
lacZ
lacY
lacA
With activator protein (crp), RNA polymerase
binds well and the transcription rate is higher.
P O
lacZ
lacY
lacA
625
3. MANY operons that encode catabolic
enzymes have the same crp binding
site ( ) and are controlled by the
same regulatory protein (CAP or crp).
crp binding site
bacterial chromosome
II. Global regulation
A. Control of many genes by a
single regulatory protein
626
operator or
activator
binding sites
of similar DNA
sequence
Bacterial chromosome
TB
B. Example: catabolite repression
A global regulatory system that
allows glucose to be consumed in
preference to a variety of other
carbon sources.
627
TB
628
1. Catabolite repression enables Escherichia
coli to use glucose in preference to other
glucose
carbon sources.
maltose
Lactose
utilization
requires
additional
proteins.
TCA
lactose
(galactose--1,4-glucose)
crp
P O
binding site
lacZ
 -galactosidase
lacY
lacA
lac permease (transport protein)
2. Key components of catabolite
repression
a. cAMP (cyclic AMP)
629
an effector molecule that increases
the DNA binding affinity of
the catabolite activator protein
b. CAP (or crp)
Catabolite activator protein, a
transcriptional regulatory protein;
also called crp (cAMP receptor protein) TB
3. CAP/cAMP binds to DNA and
regulates transcription.
CAP (or crp)
630
CAP (or crp)
binding sites
cAMP
cAMP/CAP
complex
bacterial
chromosome
TB
4. How does catabolite repression work?631
a. Genes needed for the catabolism of
many carbon and energy sources
require cAMP/CAP for expression.
*b. Glucose decreases cellular cAMP levels.
c. Without cAMP/CAP, genes required to
catabolize nonglucose energy sources
are transcribed at very low rates.
d. Therefore, glucose is preferentially used
as a carbon and energy source.
C. Global regulation is often used
together with other more specific
regulatory systems.
Example: the lactose operon
requires both lactose and
cAMP/CAP for induction.
632
633
Both lactose and cAMP/CAP are needed
for high induction of lac operon.
glucose decreases cAMP
crp
P O
binding site
P O
lacZ
lacY
glucose present,
lactose absent
lacA
lac repressor binds DNA in absence of lactose
lacZ
glucose absent,
lactose present
lacY
lacA
634
III. Two-component regulatory systems
Transcriptional regulatory systems
composed of a sensor kinase and
response regulator.
TB
A. Sensor kinase
Integral membrane proteins that
sense environmental conditions and
phosphorylate proteins
635
B. Response regulator
Cytoplasmic transcriptional regulatory
proteins controlled by sensor kinases
through phosphorylation
TB
effector
sensor
kinase
636
P
phosphorylation
dephosphorylation
response
regulator
P
P
cytoplasmic
membrane
TB
C. Transcriptional control
637
Phosphorylation changes the DNA
binding affinity of the response
regulator.
When response regulators are
bound to DNA, they induce or
repress gene expression.
TB
638
IV. Attenuation
A "fine tuning" system for regulating
gene expression by control of
transcriptional termination.
conformation 2
1
2
3
4
UUUUU
intrinsic
transcriptional
terminator
TB
639
A. The trp operon is regulated at
two levels.
1. repression by trp repressor (on/off)
2. attenuation (fine tuning by
transcriptional termination)
R
R
P O
E
D
C
B
A
genes encoding the enzymes used
for tryptophan biosynthesis
B. Leader region and leader peptide
leader
region
P O
640
tryptophan biosynthesis genes
"structural genes"
E
D
C
B
A
transcription
translation
leader
peptide
mRNA
tryptophan biosynthetic enzymes
1. leader region of trp mRNA
641
mRNA region upstream of the coding
region for the trp biosynthesis enzymes
leader coding region for trp enzymes
region
trp mRNA
TB
2. leader peptide
A short peptide encoded by the
leader region of the trp mRNA.
642
trp leader mRNA
(has 2 trp codons)
translation
leader peptide
met-lys-arg-ile-phe-val-leu-lys-gly-trp-trp-arg-thr-ser
(Don't memorize sequence)
TB
643
C. The trick to attenuation
1. The leader region of the trp mRNA
has four segments that can fold into 2
mutually exclusive conformations by
complementary base pairing.
trp mRNA leader region
1
2
3
4
TB
mRNA leader region
1
2
3
644
4
conformation 1
1
2
3
conformation 2
1
2
3
4
intrinsic
transcriptional
terminator
4
UUUUU (3:4 loop) TB
645
2. Conformation 2 of the
trp mRNA leader is
1 2 3 4
UUUUU
an intrinsic terminator.
If conformation 2 is formed, transcription
of the trp operon is terminated before
the remainder of the trp mRNA is made.
Plentiful tryptophan favors conformation
2 and termination. Energy is not wasted
making tryptophan when it is plentiful. TB
3. The rate of TRANSLATION of the 646
leader peptide determines which
conformation (stem-loop) will form.
2:3 stem loop
1 2 3
4
1
2 3
mRNA
4
Intrinsic
terminator(3:4 stem-loop)
TB
4. The leader region encodes two
tryptophans in a row.
647
a. When tryptophan is plentiful,
translation of the trp leader peptide
is FAST (i.e. ribosomes move fast).
Fast translation favors formation of
the intrinsic terminator (the 3:4 loop).
Transcription terminates before the
structural genes are transcribed.
648
b. When tryptophan levels are LOW,
translation of the trp leader peptide
is SLOW. The ribosome PAUSES at the
trp codons, waiting for tryptophan-tRNA.
When the ribosome pauses, the 2:3
stem-loop forms. The 3:4 intrinsic
terminator stem-loop CANNOT form.
Transcription of the trp biosynthesis
genes continues.
C. Transcription of the trp operon
when tryptophan is plentiful.
Ribosome begins translation
immediately after RNA
synthesis occurs.
1
1
1
649
Ribosome finishes translation
of the leader peptide and leaves
the mRNA.
2
2
Stem loop 1:2 forms.
TB
1
1
2
3
2 3
4
4
leader
region
P O
650
Transcription continues.
Stem loop 3:4 forms.
Stem loop 3:4 is an
intrinsic terminator that
prevents further
transcription.
XE XD XC XB XA
1 2 3 4
UUUUU
tryptophan biosynthesis
genes are NOT transcribed TB
D. Transcription of the trp operon
when tryptophan is low.
651
Ribosome begins translation
immediately after RNA
synthesis occurs.
1
1
2
1
2
3
Because tryptophan is low, the
ribosome pauses at tryptophan
codons of the leader peptide and
remains attached to the mRNA.
Transcription continues.
TB
1
1
2
2
The ribosome blocks
base pairing between
segments 1 and 2.
3
3
4
652
Segments 2 and 3 pair
blocking the pairing of
3 and 4.
no terminator is formed
Note that the alternative conformations of the trp
leader mRNA are mutually exclusive.
TB
End result: When tryptophan levels
are low, the genes for the tryptophan
biosynthesis are expressed.
P O
1 2
E
34
D
C
B
A
653
DNA
mRNA
proteins
no terminator is formed
tryptophan biosynthesis
genes are transcribed
and translated
Study objectives:
Please study both the concepts and details of Regulation.
654
1. What is an activator protein? How does it work? What is the catabolite
activator protein or cAMP receptor protein (crp)?
2. What are the roles of cAMP, CAP and glucose in catabolite repression?
3. What is global regulation? Describe the example presented in class.
4. How do the lac repressor system and cAMP/CAP system regulate expression
of the lac operon? Understand (in detail) the effects of both lactose and
glucose on the expression of the lac operon.
5. Describe how sensor kinases and response regulators function in
two-component regulatory systems.
6. Understand the CONCEPTS and DETAILS of attenuation.
What is the role of tryptophan, the leader peptide, the ribosome, and
alternative leader mRNA conformations in trp operon attenuation?
7. Compare and contrast (i) transcriptional regulation by regulatory
proteins (ii) two-component regulatory systems and (iii) attenuation.
655
MCB 3020, Spring 2004
Chapter 8:
Viruses
656
Viruses:
I. General properties of viruses
II. Examples of viruses
III. Viral structure
IV. Phage reproduction
V. Reproduction of lysogenic phage
VI. Overview of animal viruses
TB
Typical viruses (30-200 nm)
envelope
nucleic acid
helical
capsid
icosahedral
capsids
657
viral
specific
proteins TB
658
I. General properties of viruses
A. small (~30-200 nm)
B. non-cellular
C. replicate within host cells and
take over the host machinery
D. released from the host cell
and infect other cells
virion = extracellular state of virus.
E. often damage or kill the host TB
659
II. Some examples of viruses
A. Human wart virus
Picture
18
Icosahedral symmetry (20 regular faces)
TB
B. Tobacco mosaic virus
660
RNA virus
Helical symmetry
Picture
19
TB
C. Flu virus
661
Picture
20
enveloped virus
TB
D. Lambda virus
662
host = a bacterium
bacterial viruses are also called
bacteriophage ("bacteria eaters") or phage
III. Viral structure
A. genomes
B. capsids
C. envelopes
D. packaged enzymes
663
TB
A. Viral genomes
664
All the hereditary material of a virus
4 - 200 genes
dsDNA
ssDNA
dsRNA
ssRNA
TB
B. Viral capsids
665
Protein shell that surrounds the genome
cross-section of
icosahedral capsid
capsid
(protein coat)
genome
Protects the viral genome
Often needed for attachment to the host cells
Usually helical or icosahedral
TB
C. Viral envelopes
666
Outermost layer of enveloped viruses
Composed of host lipids and viral proteins
Often used for attachment to the host cell
lipids from host
viral proteins
TB
667
D. Packaged proteins
Proteins found within the capsid
Different functions in different viruses
viral protein
e.g. reverse transcriptase
RNA-dependent RNA polymerase
TB
1. Reverse transcriptase
668
enzyme that synthesizes DNA from
an RNA template
2. RNA-dependent RNA polymerase
enzyme that synthesizes RNA from
an RNA template
TB
IV. Reproduction of phage
A. Attachment
B. Penetration
C. Expression of viral genes
D. Genome replication
E. Capsid formation
F. Packaging
G. Release
669
TB
A. Attachment
670
Binding of a capsid or envelope
protein to a host receptor.
host receptor
(usually a specific protein,
lipid, or polysaccharide)
host cell
Specificity for the host receptor
determines virus host range
TB
Attachment and penetration
671
Virus tail fibers interacting with core polysaccharides
672
B. penetration
injection of
viral nucleic acid
and packaged
proteins.
TB
C. Expression of viral genes
673
viral genome
host machinery
Viral proteins
TB
Typical viral proteins
674
capsid proteins
proteins that block host gene expression
proteins that block restriction systems
proteins for genome replication
proteins for assembly of viral particles
TB
D. Genome replication
various methods: for example,
host enzymes only
viral enzymes only
host and viral enzymes
675
TB
E. capsid formation
self-assembly of capsid proteins
676
TB
F. packaging
677
Insertion of the nucleic acid into
the capsid
Method varies
The "headfull" method is common
TB
G. Release
1. Lysis
678
TB
2. Budding (enveloped viruses)
679
host lipids
viral
proteins
TB
V. Reproduction of lysogenic phage
A. lysis
B. lysogeny
C. prophage induction
680
TB
A. Lysis
The most frequent method of
reproduction
Occurs as described above
681
TB
B. Lysogeny
1. Prophage integration
682
bacterial
chromosome
integration
prophage
(integrated virus)
lysogen
(cell with integrated virus)
TB
2. Prophage replication
683
host replication
TB
C. Prophage induction
684
Excision of the prophage
followed by lytic replication.
UV light and other DNA damaging
agents cause prophage induction.
TB
VI. Overview of animal viruses
1. Attachment and penetration
binding to host receptor
and uptake by endocytosis
uncoating
animal cell
685
TB
686
2. Gene expression and genome
replication for animal viruses must
follow (or adapt to) eukaryotic rules
eukaryotic RNA processing
compartmentation
(nucleus vs. cytoplasm)
What features of transcription and
translation would differ between phage
and animal viruses?
TB
B. Host interactions
1. lysis
2. persistent infection
3. latent infection
4. transformation
687
TB
1. lysis
688
destruction of the host cell
2. persistent infection
viruses bud from host over a long
period of time.
3. latent infection
infections that reoccur periodically
4. transformation
increased growth rate of host cells TB
Study objectives
689
Please understand ALL the CONCEPTS and DETAILS presented in this lecture.
1. Describe the general properties of viruses.
2. Define virions, bacteriophage, phage.
3. Describe viral genomes, capsids (protein coats or shells), envelopes,
and packaged proteins. What are the functions of these molecules?
Know the specific examples presented in class.
4. Compare and contrast the details of the reproductive cycles of phage
and animal viruses. Thought question: What features of transcription
and translation would differ between phage and animal viruses?
5. How do phage reproduce by lysogeny?
6. What is a lysogen? a prophage?
7. What effects can animal viruses have on their hosts?
690
Eukaryotic viruses, viroids, and prions:
I. Polio virus
II. Flu virus
III. HIV virus
IV. HIV replication
V. HIV treatment
VI. Viroids
VII. Prions
I. Polio virus
A. Basic properties
+ssRNA
icosahedral
nonenveloped
infects nerve cells
691
+ RNA (plus strand RNA)
means that the RNA genome reads
the same as the mRNA
mRNA
5'
3'
C AA
GGUUC
+ RNA
5'
G G U U C C A A 3'
692
B. Life cycle
1. penetration and uncoating
693
nucleus of cell
uncoating
+ssRNA
nerve cell
cytoplasm
2. Genome replication
+RNA (genome)
viral RNA-dependent
RNA polymerase
-RNA
694
3. Gene expression
+RNA (mRNA)
695
polio
genome
translation (host machinery)
polyprotein
auto-proteolysis and proteolysis
coat proteins, proteases, RNA polymerase etc.
a. Gene expression facts
Polio mRNA can be translated
without a eukaryotic 5' cap
(methylguanosine cap).
Polio inactivates translation of
host mRNAs by destroying the
host protein that recognizes the
methylguanosine cap.
696
4. Assembly and release
697
+ strand RNAs are assembled into
capsids and the host cell is lysed.
II. Flu virus
698
A. Basic properties
-ssRNA
segmented genome
enveloped
helical capsid
infects mucus membrane cells
of the respiratory tract
B. Structure
viral envelope
699
hemagglutinin
neuraminidase
segmented genome
(-RNA)
700
- RNA (minus strand RNA)
is complementary to the mRNA
mRNA
5'
3'
C AA
GGUUC
- RNA
3'
C CAA G G U U
5'
C. Key proteins
701
1. Hemagglutinin
mediates fusion of the viral
envelope to the host cell membrane
2. Neuraminidase
Breaks down sialic acid
and assists in budding
D. Antigenic shift
Major changes in viral proteins due
to mixing of genome segments from
different viruses.
Occurs when two different viruses
infect the same host.
This can cause dramatic changes
in surface antigens and produce
new virulent strains.
702
III. HIV (AIDS) virus
703
Human immunodeficiency virus
HIV kills CD4+ cells of the immune system
Causes AIDS
Healthy adults have about 800 CD4+
T-cells/cubic millimeter of blood.
HIV patients are said to have AIDS when
they develop opportunistic infections or when
their CD4+ T-cell count falls below 200.
A. HIV infection
704
Usually acquired by sexual intercourse
Almost always fatal
No cure
No vaccine
In the US,~1/250 people are infected.
In the US,~1/3,000 people contract HIV each year
On average, 8-10 years pass between
HIV infection and the development of AIDS.
B. Prevention
705
Celibacy
Insistence on condoms
Clean needles
Post-exposure drug treatment within 24 h ???
4-weeks of treatment with possible side
effects of headache, nausea, fatigue
and anemia.
C. HIV replication
HIV is a retrovirus = an RNA
virus that replicates through
a DNA intermediate.
reverse
transcriptase
HIV genome
+ ssRNA
(2 copies)
DNA
706
D. HIV structure
707
envelope protein
reverse transcriptase
integrase
protease
+ssRNA
Genetic map of typical retrovirus
gag
pol
LTR = long terminal repeat
env
other
genes
708
LTR
gag: encodes internal structural proteins
pol: encodes reverse transcriptase
env: encodes envelope proteins
There are also other genes specific to
different retroviruses.
A. HIV proteins
709
1. Envelope protein:
mediates binding to CD4 receptor
2. Reverse transcriptase:
synthesizes DNA from an
RNA template
3. Integrase:
splices viral DNA into the
host genome
4. Protease
cleaves the viral polyprotein
into active parts
710
E. HIV reproductive cycle
711
g
a
b
f
CD4 receptor
c
cell membrane
nuclear
membrane
d
e
HIV provirus
712
Steps in the HIV reproductive cycle
a. penetration and uncoating
b. reverse transcription
c. integration
d. gene expression
e. replication
f. polyprotein cleavage by HIV protease
g. assembly and budding
713
F. HIV Treatment
A. Reverse transcriptase inhibitors
B. protease inhibitors
In general, two reverse transcriptase
inhibitors are used in combination
with a protease inhibitor; however,
treatment is complex and rapidly changing.
http://www.hivatis.org/trtgdlns.html
(The latest information on HIV treatment)
G. HIV drug resistance
protease
inhibitor
HIV protease
mutation
drug
resistant
protease
714
inhibitor binding
to the active site
inactivates the
protease
inhibitor no
longer binds but
protease still
functions
VI. Viroids
715
circular single stranded RNA
molecules that cause plant diseases
viroids are "naked" RNA
(no proteins associated with RNA)
viroid genomes do NOT
encode proteins
VII. Prions
Infectious proteins
Prion proteins appear to transmit
disease without DNA or RNA.
716
A. Prion diseases
(spongiform encephalopathies)
Scrapie, sheep and goats
Mad cow disease, cows
Creutzfeldt-Jacob, humans
717
Mad cow disease (BSE)
• Bovine spongiform encephalopathy (BSE)
• source of infection appears to be feeding
cows with "meat-and-bone meal" remains
of infected sheep or cows, especially
infected brain tissue
• prion is not destroyed by cooking
718
719
"new variant" Creutzfeldt-Jacob syndrome
• human disease thought to be caused by eating
BSE-infected beef
• about 92 cases, most victims have died
• unusual in that many victims are < 30 years old
• incubation time is 10 to 15 years
B. How prions cause disease
720
normal PrP (prion protein)
Disease-causing PrP catalyzes
a conformational change
that turns normal PrP into
disease causing PrP.
disease causing PrP
Over time, disease causing PrP accumulates and
symptoms result.
C. The prion gene
If a protein transmits the disease,
where is its gene?
The prion gene (prp) turned out to be a
normal gene found in animals.
Unusual forms of the gene (mutants)
are thought to cause disease.
721
Study objectives
722
1. Describe the structure of the polio virus. Explain polio virus replication
and gene expression. What are polyproteins?
2. Distinguish between plus strand and minus strand RNA genomes.
3. Describe the structure of the flu virus. What is the relationship of flu virus
genome structure to antigenic shift.
4. What are the functions of hemagglutinin and neuraminidase.
5. How is HIV transmitted?
6. How is HIV infection prevented?
7. How is HIV infection treated?
8. Describe the structure of HIV. What is a retrovirus? Describe the
general structure of a retroviral genome and the proteins encoded.
9. Describe the HIV reproductive cycle. Know the functions of the HIV proteins.
10. What are viroids? How do viroids differ from viruses and prions?
11. What are prions?
12. What diseases do prions cause?
13. How are prions thought to cause disease?
14. Where are prions genes found?