all cells...
A. have membrane transport proteins.
B. synthesize proteins on the ribosome.
C. replicate their genome by DNA polymerization.
D. transcribe their genetic information by RNA polymerization.
E. All of the above.
E
Which of the following would you NOT expect to find in a bacterial cell?
A. Swimming using flagella
B. Having a cell wall around the plasma membrane
C. ATP production in mitochondria
D. Protein production on the ribosome
E. Sexual exchange of DNA with oth
C
You run a mixture of proteins on an SDS-PAGE and identify two proteins: Cyclin-B and Ribonucleotide reductase. From the position on the gel you estimate that these proteins are about 57 kDa and 44 kDa respectively.
The predicted molecular masses based on
C - slight difference can be caused by composition of the amino acid (B?!)
Protein secondary structure elements such as ? helices and ? sheets constitute the major regular folding patterns in proteins. With regard to these elements, ...
A. hydrogen-bonding between the amino acid side chains defines the type of secondary structur
D
Which statement about Chaperones is NOT true?
A. Most proteins need the assistance of Chaperones to fold inside the cell.
B. Some proteins are folded only by HSP-60 and do not need HSP-70
C. HSP-70 binds to proteins as they are being synthesized by the ri
B
How do Chaperones recognize proteins that are NOT completely folded? (so they can bind to those proteins to help them fold)
A. Chaperones can recognize the shape of unfolded proteins.
B. Chaperones can recognize the shape of folded proteins.
C. Chaperones
C
Which of the following proteins determines which protein lives or dies by ubiquitin-mediated degradation?
A. HSP-70
B. Ubiquitin Activating Enzyme
C. E2
D. E3
E. Proteasome core
D
You have a drug that blocks the activity of one of the enzymes involved in ubiquitylation of proteins. You treat cells with this drug and you determine the effect of treatment on the abundance of different proteins by Western blot analysis. Cdc2 is a cont
A
You have a drug that blocks the activity of the Proteasome. You treat cells with this drug and you determine the effect of treatment on the abundance of your favorite protein by Western blot analysis.
The arrow points to the size of your favorite protein
E
Chromosome 3 contains nearly 200 million nucleotide pairs of our genome. If this DNA molecule could be laid end to end, how long would it be? The distance between neighboring base pairs in DNA is typically around 0.34 nm.
A. About 7 mm
B. About 7 cm
C. Ab
D?
Stable ?-sheet aggregates can form from many proteins, forming intertwined cross-beta strands that have the potential to kill cells or damage tissues. Which of the following is NOT true regarding these aggregates?
A. They form almost exclusively in the ce
A
On the ribosome, the mRNA is read from ..., and the polypeptide chain is synthesized from...
A. 5' to 3'; C- to N-terminus.
B. 5' to 3'; N- to C-terminus.
C. 3' to 5'; C- to N-terminus.
D. 3' to 5'; N- to C-terminus.
B
DNA and RNA polymerase differ in all of the following EXCEPT...
A. the nucleotide substrates they incorporate.
B. their requirement for a primer.
C. their error rate.
D. the type of chemical reaction they catalyze.
E. their processivity.
D
How does a eukaryotic cell deal with the superhelical tension in its genomic DNA resulting from the activity of RNA polymerases?
A. DNA gyrase introduces negative supercoils, keeping the DNA under constant tension.
B. The RNA polymerases are allowed to ro
C
Eukaryotic pre-mRNAs undergo a number of modifications such as capping at the 5' end. A 5' cap...
A. consists of a modified terminal adenine nucleotide.
B. has a 3'-to-5' linkage between the terminal nucleotide and the 5' end of the pre-mRNA.
C. contains
C
Stable ?-sheet aggregates can form from many proteins, forming intertwined cross-beta strands that have the potential to kill cells or damage tissues. Which of the following is NOT true regarding these aggregates?
A. They form almost exclusively in the ce
A
You are studying a protein of 100 kilodaltons (kd). You found a way to cleave this protein at a single site � of the length from the N-terminus. You have raised a monoclonal antibody that binds to the N-terminus of protein X.
You run on an SDS-PAGE a samp
D
Polysomes...
A. are large cytoplasmic assemblies made of several ribosomes each translating their exclusive mRNA.
B. are only found in the eukaryotic cytoplasm.
C. can take advantage of the circularization of eukaryotic mRNA (by interactions between the 5
C
R group of serine
CH2 OH
general amino acid structure
H2N - CH - COOH (+R group)
polar neutral amino acids
found at surface of proteins
mostly CH side chains
COOH groups have low enough pka's to lose protons
3 points of 1800's Cell Theory:
1) a cell is the structural unit of life
2) all organisms are made of cells
3) all cells arise from pre-existing cells
cell theory was formulated in ___ by ____ and _____
1838
Robert Hooke
S. Schleiden
4 things common to prokaryotes and eukaryotes:
DNA
plasma membrane
cytoplasm
ribosomes
a cell consists mostly of:
water and proteins
forming bonds ____ energy, while breaking bonds _____ energy
releases, requires
in hydroxyl water bonds, the O is partially (+/-) and the H is partially (+/-)
O is negative
H is positive
molecules without _________ tend to be less ? reactive
electronegative atoms
(waxes and fats, ex.)
polar ionization
atoms that are so strongly electronegative they can capture electrons during chemical reactions
ex= Na + Cl- (relatively stable)
noncovalent bonds
interactions between molecules or different parts of large molecules
typically weaker bonds, but depends on attractive forces between + and - parts of molecule(s)
types of noncovalent bonds (4)
ionic
hydrogen bonding
van der Waals forces
hydrophobic forces
ionic bonds are strong in ___ but cannot hold attraction in ____
crystal, water
ionic bonds in a cell are generally:
weak due to the water in the cell
(stronger in protein core)
hydrogen bonds
hydrophilic
enhance solubility in and interactions with water
noncovalent bond
hydrogen bonds occur between most ___ molecules
polar
(also between polar groups and large biological molecules like DNA)
hydrogen bonds are: (strength)
strong collectively, weak individually
2-5 kcal/mol in water
van der Waals forces
hydrophobic groups can form weak bonds with one another based on electrostatic interactions;
due to slight perturbations of electron distributions
dipoles
transient charge asymmetries resulting in momentary charge separations
van der Waals forces: (strength)
very weak as single forces
0.1-0.3 kcal/mol
sensitive to distance
must be close and of complementary shapes for force
van der Waals forces play important roles in:
antibody - antigen interactions
molecules with nonpolar covalent bonds lack _____ that can _____ and are thus ______
charged regions
interact with polar water molecules
insoluble
hydrophobic forces
molecules with nonpolar covalent bonds form into aggregates to minimize exposure to polar molecules/solutions
peptide bonds are:
covalent
disulfide bridge bonds are:
covalent
primary protein structure
amino acid sequence joined by peptide bonds
secondary protein structure
local structure formed through noncovalent H-bonds between amino acids over short stretches of a polypeptide chain
tertiary protein structure
the overall shape of a polypeptide chain formed through non-covalent bonds between amino acids over long stretches of a polypeptide chain
quaternary protein structure
formed when 2 or more polypeptide chains specifically associate with one another to form a protein complex
Glycine
G
non polar
Alanine
A
non polar
Valine
V
non polar
Proline
P
non polar
Leucine
L
non polar
Isoleucine
I
non polar
Methionine
M
non polar
Tryptophan
W
non polar
Phenylalanine
F
non polar
Cysteine
C
non polar (- when deprotonated)
Tyrosine
Y
polar (- when deprotonated)
Serine
S
polar
Threonine
T
polar
Asparagine
N
polar
Glutamine
Q
polar
Lysine
K
positive
Histidine
H
positive
Arginine
R
positive
Aspartic Acid
D
negative
Glutamic Acid
E
negative
alpha helix
right hand coil
every backbone N-H group is hydrogen bonded to the C=O group of the 4th amino acid residue
example of quaternary structure
alpha-helical coiled-coil
formed by 2 peptides twisting around each other and stabilized by hydrophobic interactions between them
quaternary = adds disulfide bonds
quaternary structure folds the protein:
into lowest energy conformation
_____ help stabilize tertiary and quaternary structures
disulfide bonds
Disulfide bonds:
single covalent bond between sulfur atoms; usually formed from the oxidation of sulfhydryl (-SH) groups
the only way to break a disulfide bond is:
to reduce it harshly with a detergent
SDS-PAGE stands for:
sodium dodecyl sulfate
polyacrylamide gel electrophoresis
sodium dodecyl sulfate
a detergent that can dissolve hydrophobic molecules but also has a negative charge (sulf-ate) attached to it
binds to hydrophobic protein regions
SDS disrupts _____ structure and gives all proteins a constant _______
secondary
charge:mass ratio
= 1.2 g SDS per g protein
proteins are separated/fractionated by _______ during gel electrophoresis
size (SDS removes charge differences)
protein phosphorylation
reaction that transfers a phosphate group from ATP to an amino acid side chain of the targeted protein via a protein kinase
removal of the phosphate group is catalyzed by a phosphatase
Protein Function: Catalysis
enzymes (hydrolitic, proteases, polymerases, kinases)
Protein Function: Defense
Immunoglobins (antibodies)
Toxins (venom)
Cell surface antigens (MHC proteins)
Circulating transporters (hemoglobin, myoglobin)
Membrane transporters (sodium-potassium pump)
Protein Function: Support
fibers (collagen, keratin, fibrin)
Protein Function: Motion
muscle (actin, myosin)
Protein Function: Regulation
osmotic proteins
gene regulators
hormones
Protein Function: Storage
ion binding
goal of a Western Blot:
obtain and purify a protein to use as an antigen
antibody
Y-shaped protein produced mainly by plasma cells that is used by the immune system to identify and neutralize pathogens
Monoclonal antibodies
monospecific antibodies that are made by identical (one lineage of) immune cells that are all clones of a unique parent cell
Polyclonal antibodies
antibodies secreted by different B cell lineages within the body; a collection of immunoglobulin molecules that react against a specific antigen and each identify a different epitope
_____ antibodies are better because ______
monoclonal
more specific
______ is the final system to control protein concentration in cells
protein degradation
most cytosolic proteins are degraded in the:
proteasome
proteins fold from ___ terminal to ___ terminal
N to C
folding of a protein is controlled by its:
amino acid sequence
heat shock proteins
proteins involved in the folding of new proteins or re-folding of proteins that started to unfold after heat treatment
both HSP70 and HSP60 use ____ to fold proteins
ATP
HSP-70 folding of proteins:
binds to hydrophobic region
many monomers bind along length of protein
can bind as soon as protein is out of ribosome
can refold a partially misfoled protein
useful to maintain a protein in an extended conformation >
HSP-60 folding of proteins:
HSP-60 is a large complex with two folding chambers
works with 70, which delivers unfolded protein
_____ degrades proteins that fail to fold so that _____
proteasome
they do not aggregate
proteasome
degrades proteins tagged by ubiquitin chains
consists of a cap and core
proteasome cap
called regulatory particle
selectively binds proteins that have been marked by ubiquitin for destruction
contains unfoldase to denature/unfold proteins using ATP hydrolysis > feeds them to core
proteasome core
unfolded proteins are fed through a narrow channel into the inner chamber of 20S cylinder for digestion
the proteasome cap removes ____ for recycling before feeding targeted protein to the core
ubiquitin subunits
ubiquitin
universal"
76 amino acid long chain -> formed as polyubiquitin chain and cut into single ubiquitins after
has hydrophobic globular core
COOH is the point of attachment to lysine side chains
E1
ubiquitin activating enzyme
the C terminal of ubiquitin links to E1
E2
ubiquitin conjugating enzyme
E3
ubiquitin ligase
functions to recognize different substrates
mono- ubiquitin
histone regulation
multi- ubiquitin
endocytosis
poly- ubiquitin
proteasomal degradation, DNA repair
Ubiquitylation can be controlled by: (2)
1) activation of a ubiquitin ligase
2) activation of a degradation signal on the target protein
who discovered DNA was the genetic material?
when?
where?
Watson and Crick
1953
Cambridge University
the structure of DNA is:
non-static
direction of DNA synthesis
5'-3'
direction of DNA polymerase proofreading
3'-5' exonuclease activity
DNA replication follows a ___ model:
semiconservative > 2 strands separate, each is used as a template to produce a complementary strand > each new DNA helix has one old and one new strand
DNA replication is continuous on:
3'-5' template (leading)
DNA replication is discontinuous on:
5'-3' template (lagging)
histone core
H2A
H2B
H3
H4
the maternal and paternal chromosomes of a pair are called
homologous chromosomes
homologs
the only non-homologous chromosome pair in humans
sex chromosomes in males - Y from father and X from mother
what percent of our genome is highly conserved?
5%
what percent of our genome is made of various repeated sequences?
50%
mostly transposable DNA elements
RNA
linear polymer made of 4 nucleotide subunits linked by phosphodiester bonds
(RNA/DNA) is more flexible
RNA
transcription
produces RNA using one strand of DNA as the template molecule
begins with unwinding of small part of DNA
RNA poly transcribes DNA by catalyzing:
the formation of phosphodiester bonds between nucleotides (5'-3')
RNA poly vs DNA poly
Both: proofreading
RNA: addition of ribonucleotides; 50/sec
does not need a primer
DNA: addition of deoxyribonucleotides (1000/s)
more accurate
mRNA
RNA copied from genes that ultimately directs synthesis of proteins (3-5% of total)
tRNA, sRNA, rRNA
final product of genes (RNA itself) (majority of total)
promoter
DNA sequence RNA polymerase binds to to initiate RNA synthesis in prokaryotes
consists of 2 short sequences at -10 and -35 positions upstream from start site
sigma subunit
RNA poly subunit that binds promoter and opens helix
dissociates after synthesis of ~10 bases
associates with core enzyme and assists in reading DNA signals for where to start transcription
prokaryote termination signal
hairpin loop and A-T string
-10 sequence
Pribnow box; essential; usually TATAAT
-35 sequence
consists of TTGACA; allows a very high transcription rate
steps of prokaryotic transcription (7)
1) RNA poly holoenzyme assembles and then locates a DNA promoter sequence
2) RNA poly unwinds and opens DNA at starting position
3) initial RNA synthesis (determining step)
4) releases sigma factor; poly tightens around DNA
5) elongation
6) termination ha
____ RNA can contain instructions for several different proteins, but ____ mRNA has information for only one protein
bacterial
eukaryotic
RNA Poly I (eukaryotic)
transcribes tRNA and rRNA and snRNA
RNA Poly II (eukaryotic)
transcribes genes encoding proteins
RNA Poly III
transcribes tRNA and rRNA and snRNA
eukaryotic promoters
lie much further away from start site > transcriptional complexes can cause DNA to bend back on itself to allow placement of regulatory sequences
contain TATA box
TATA box
TATAAA
binds to a TATA binding protein which assists in formation of RNA poly transcriptional complex
normally very close to start site
RNA poly II requires ___ general transcription factors
5
basal level requires:
basal transcription factors to bind DNA
activator
eukaryotes
bind to specific sequences and facilitate binding of GTF's and RNA poly
mediator
eukaryotes
enable activators to interact with GTF's and RNA poly
chromatin-modifying proteins
eukaryotes
remodeling complexes allow greater accessibility to DNA
elongation factors
ensure RNA poly does not associate before end of the gene is reached; associated with RNA poly shorty after transcription initiation
DNA topoisomerases
remove superhelical tension
DNA gyrases
use ATP to pump supercoils into DNA
RNA processing steps: (3)
1) eukaryotic mRNA capped at 5' end
2) polyadenylated at 3' end (poly A tail)
3) introns removed
phosphorylation of RNA tail CTD consists of ___ and promotes ____ and allows ____
consists of domain of 7 amino acids (containing 2 phosphorylated serines) repeated 52 times
promotes disassociation of RNA poly from proteins present at start of transcription
allows new set of elongation proteins and pre-mRNA processing proteins to assoc
capping of pre-mRNA's with modified ______ occurs after ~ ____ bases
guanine nucleotide (G)
25
capping of pre-mRNA's involves 3 enzymes:
phosphatase - removes one p' from 5' end of mRNA
guanyl transferase - adds GMP to 5' end
methyl transferase - adds methyl group to guanosine
splicing machinery recognizes 3 portions (conserved) of the pre mRNA:
5' splice site
3' splice site
branch point in the intron that forms the base of the excised lariat
steps of RNA splicing mechanism (9):
1) spliceosome recognizes splicing signals on pre-mRNA, brings ends of intron together
2) branch point site recognized by BBP and U2AF
3) U2 snRNP displaces BBP base pairing with branch point consensus sequence
4) U1 snRNP base pairs with 5' splice site j
the spliceosome's mediation of splicing RNA is performed primarily by:
the 5 snRNA molecules (U1, U2, U4, U5 (U6?)) forming the spliceosome core
___ required to break RNA-RNA interactions
RNA helicase requiring ATP
all steps in splicing except _____(2) required ATP and other proteins
association of BBP with branch chain A
and U2 with 5' splice site
RNA splicing has:
plasticity and flexibility - enables cell to regulate pattern of RNA splicing
splicing patterns are regulated so that:
different forms of protein produced at different times and in different tissues
Group I intron (self-splicing intron)
reactive G nucleotide attacks the initial phosphodiester bond cleaved during the splicing reaction
Group II intron (self-splicing intron)
reactive A in intron sequence is attaching group, and lariat intermediate generated
CstF and CPSF proteins:
associate with RNA poly tail transferred to RNA as it emerges
CstF = cleavage stimulating factor F
CPSF = cleavage processing specificity factor
export of mature mRNA's from nucleus is:
highly selective and coupled to correct mRNA processing
mRNA is only exported if appropriate set of proteins are bound, including:
cap binding complex
snRNP proteins absent
proteins that mark complete splicing
hnRNP's
heterogeneous nuclear ribonuclear proteins; =most abundant proteins that assemble on pre-mRNA as it emerges from RNA poly
most of the RNA in the cell:
performs a catalytic or structural function
80% of total RNA is
rRNA
=transcribed by RNA poly I (which has no C terminal tail); neither capped nor polyadenylated
3-5% of total RNA is
mRNA
the nucleolus is a site for:
processing rRNA's and assembling them into ribosomes
the size varies and reflects the # of ribosomes the cell is making
also a site where other RNA's are produced and other RNA-protein complexes assembled (tRNA's, snRNA's, U6, telomerase...)
alternative splicing
alternatively selecting reading frames
ribosomes are formed by
proteins and RNA's
ribosomes consist of:
a large subunit (3 tRNA binding sites = A, P, E) and a small subunit (mRNA binding site)
[aminoacyl, peptidyl, exit]
ribosomes can be found
either freely floating in the cytosol or bound to the rough ER (in eukaryotes)
transfer RNA folds into a _____ with the 4 key regions of:
cloverleaf structure
1) acceptor stem = 3' CCA that carries an amino acid
2) anticodon = associated with the mRNA codon (complementary)
3) T arm = associates with the ribosome via A,P,E sites
4) D arm = associates with the tRNA activating enzyme (adds the
the first stage of translation is ____ and involves:
initiation
the assembly of the 3 components that carry it out (mRNA, tRNA, ribosome)
if RNA doesn't have a cap:
it will be degraded
polysome
a group of 2 or more ribosomes translating an mRNA sequence simultaneously
difference in prokaryotes regarding polysomes?
they may form while the mRNA is still being transcribed from the DNA template
ribosomes located at the ___ end of the polysome cluster will have longer polypeptide chains than those at the ___ end
3'
5'
all proteins begin being synthesized on:
ribosomes in the cytosol (except for few being synthesized on mitochondria or plastid ribosomes)
Golgi apparatus
functions as a molecular assembly line in which membrane proteins undergo extensive post-translational modification
Lysosomes
break down macromolecules into their constituent parts, which are then recycled
lysosomes contain enzymes called ____ that _____. They are called ____ in plants and fungi.
hydrolases that can digest proteins, nucleic acids, lipids, and complex sugars
acidic vacuoles
lysosomes are formed by the fusion of:
vesicles that have budded off from the trans-Golgi