EXAM 2 Flashcards
1
Q
formula of carb
A
Cn(H2O)n
2
Q
how are carbs produced
A
from Co2 and H2O via photosynthesis
3
Q
small carb
A
glyceraldehyde (90 g/mol)
4
Q
large carb
A
amylopectin (200million g/mol)
5
Q
functions of carbs
A
energy source and storage
structural component of cell walls and exoskeletons
informational molecules in cell-cell signaling
6
Q
configuration of a monosaccharide is based on the OH on the
A
penultimate carbon
7
Q
epimer
A
diastereomers that differ only in configuration of 1 carbon
8
Q
D-fructose
A
ketose of glucose at C2
9
Q
D-galactose
A
C4 epimer of glucose
10
Q
D-mannose
A
C2 epimer of glucose
11
Q
most common monosaccharides
A
hexoses
D-glucose
D-fructose
pentose
D-ribose
12
Q
aldose + mild oxidizing agent =
A
aldonic acids
13
Q
aldoses are
A
reducing sugars
14
Q
naturally occurring ketoses almost always carry the keto group on
A
carbon 2
15
Q
aldehyde and ketone carbons are
A
electrophilic
16
Q
reactions of aldehyde/ketone and alcohol oxygens allow monosaccharides to form
A
ketones
17
Q
when 2 monosaccharides are bound together
A
acetals and ketals form
18
Q
monosaccharides with sufficiently long chains can cyclicize spontaneously… leads to
A
pyranose or furanose
19
Q
pyranose
A
6 carbons
20
Q
furanose
A
5 carbons
21
Q
how do cells prevent formation of fructopyranose?
A
glucose and fructose are phosphorylated at C6
22
Q
how does carbon 6 point for a D isomer
A
up
23
Q
how does hydroxyl on animeric carbon point
A
right is down
24
Q
animeric carbon
A
new chiral carbon in haworth perspective formula
25
fructofuranose has ____ not in the ring
C1 and C6
26
in glucose chair conformations
all OH groups are equatorial except C1, the anomeric carbon in alpha form
27
ribose in nucleotides is usually
phosphorylated at C5
28
lactose connection
B1-4
29
lactose molecules
B galactose + B glucose
30
sucrose connection
a1, 2B
31
sucrose molecules
B fructose + a glucose
32
maltose molecules
2 glucose
33
what connects two monosaccharides into disaccharides?
glycosidic bond
34
what is produced by most mammals but not us
galactose - a-1,3 - galactose
can acquire allergy by bite of lone star tick or chemo from mice
35
polysaccharides
natural carbohydrates, usually found as polymers for storage
36
polysaccharides can be
linear
branched
homopolysacc
heteropolysacc
37
how are polysaccharides made?
no template -- made based on concentrations of monosaccharides
38
polysaccharides molecular weight
not defined
39
glycogen
branched homo-disccharide of glucose
| MW reaches several millions
40
glycogen structure
glucose monomers form a1-4 chains
| branch points with a1-6 linkages every 8-12 residues
41
glycogen function
main storage polysaccharide in animals
42
starch
mixture of 2 homopolysaccharides of glucose
main storage polysaccharide in plants
amylose
amylopectin
43
amylose
starch; long unbranched polymer of a1-4 linked residues
44
amylopectin
starch; branched (like glycogen) branch points with a1-6 linkages every 24-30 residues
45
cellulose
unbranched homopolysaccharide of glucose
tough, water insoluble
most abundant polysacch, found in plant cell walls
46
cellulose structure
glucose monomers form b1-4 linked chains
hydrogen bonds form between adjacent monomers
additional H bonds between other cellulose chains
47
cellulose metabolism
difficult to digest
cellulase allows fungi, bacteria, protozoa to use wood as glucose
ruminants and termites live symbiotically with microorganisms that secrete cellulase
48
why is cellulose difficult to digest?
fibrous structure and water insolubility --> not many enzymes hydrolyze b1-4 linkages
49
chitin
major building block in exoskeleton of arthropods
cell walls in some fungi
tough, flexible, water insoluble (more than cellulose)
50
chitin structure
identical to cellulose but monomer is N-acetylglucosamine
51
tertiary structure of polysaccharides is determined by
weak interactions
regions of rigid structures
steric hinderance
limited bonds that can rotate
52
tertiary structure of starch ang lycogen
form tightly coiled helix
| stabilized by H bonds
53
tertiary structure of cellulose
extended structure with H bonding between chains
54
glycoproteins
proteins with oligosaccharides attached
55
glycoprotein structure
anomeric carbon binds to the serine or threonine residues (O linked) or the asparagine (N linked)
56
how many proteins in mammals are glycosylated?
1/2
57
how does adding oligosaccharides to proteins affect the protein?
increases solubility
can target proteins for certain cellular locations
can change protein structure
58
Nucleotide functions
building blocks
```
energy for metabolism (ATP)
enzyme cofactors (NAD+)
signal transduction (cAMP)
```
59
nucleic acid functions
storage of genetic info (DNA)
transmission of genetic info (mRNA)
processing of genetic info (ribozymes)
protein synthesis (tRNA, rRNA)
60
Nucleoside structure
no phosphate
61
phosphate group
negatively charged
on 5' carbon of pentose ring
(sometimes other positions)
62
nucleic acids are built using
5'-triphosphates (ATP, GTP)
63
nucleic acid structure
one phosphate group per nucleotide molecule
64
nucleotide structure
can be 5'diphosphate, 5'-triphosphate
65
C2' Endo
C2 carbon off plane, 5' carbon on the same plane
66
C3' Endo
C3' carbon off of plane, 5' carbon on same plane
67
nitrogenous bases
derivatives of pyramidine or purine
nitrogen containing heteroaromatic molecules
planar or almost planar
68
pyrimidine bases
```
1 ring
cytosine
thymine
uracil
all good G bond donors and acceptors
```
69
purine bases
2 rings
adenine
guanine
good H bond donor and receptors
70
B-N-glycosidic bond
anomeric carbon in b conformation
N1 in pyrimidines
N9 in purines
difficult to hydrolyze
71
UV abs at 250-270
used to determine concentration of a solution of nucleic acids
72
conformation of N glycosidic bond
can rotate with N-C1 (chi)
0* = syn
180 * = anti
73
purine conformation
syn, anti
74
pyrimidine conformation
anti only because interactions of sugar ring and =O
75
B dna conformation
anti conformation
76
minor nucleosides in DNA
5 methylcytidine
N6 methyladenosine
77
5 methylcytidine
eukaryotes and some bacteria
78
N6 menthyladenosine
bacteria not eukaryotes
79
epigenetic marker
phenotype change without change in DNA
80
prokaryotic epigenetic markers
mark own DNA so only foreign DNA is degraded
81
eukaryotic epigenetic markers
marks which genes should be active
methylated = not transcribed
82
nucleotide functions
energy source
coenzymes
regulatory molecules
83
nucleotide functions: energy source
3 PO4 provides energy by cleaving
84
nucleotide functions: coenzymes
Coenzyme A --> nucleotide makes up part of it, not prosthetic group
85
nucleotide functions: regulatory molecules
cAMP, cGMP = signaling molecules part of signal transduction and amplification
86
oligonucleotides
phosphodiester bonds link nucleotides covalently
PO4 from C5' onto C3 oh of another
negatively charged backbone
linear polymers
directionality
87
DNA backbone
fairly stable
88
RNA backbone
less stable
OH at C2' can bind to PO4, breaking covalent bond at C5' of next nucltodie
cyclic PO4 between C2' and C3'
89
H bonding in oligonucleotides in RNA
G pairs with U for structure formation or codon/anticodon wobble
90
base stacking in oligonucleotides
```
holds helix together
noncovalent interactions (hydrophobic, van der waals)
```
91
franklin and wilkins
cross in x ray crystalography = helix
diamonds = sugar po4 backbone on outside
calculated helical parameters
92
watson and crick
missing layer in cross = major and minor groove
H bonding AT/CG
double helix
93
Forms of DNA
A
B
Z
94
A DNA
wider, c3' endo (more like RNA)
all anti
Franklin found it in dehydrated DNA
95
B DNA
C2' endo
all anti
Watson and crick
96
Z DNA
left handed helix
pyrimidines: c2' endo, anti
purines: c3' endo, syn
may be formed during transcription
97
DNA denaturation
separates 2 strands of DNA
covalent bonds remain in tact, genetic code remains in tact
UV ABS INCREASES due to loss of interactions between bases
high temp or change in pH
reversible, can bind other molecules
98
Factors affecting Tm
base composition
DNA length
ionic strength
99
Tm
midpoint of melting
100
Factors affecting Tm: base composition
more CG = increases Tm
101
Factors affecting Tm: DNA length
longer DNA increases Tm
more interactions = stronger base stacking
102
Factors affecting Tm: Ionic strength
high salt increases Tm
salt binds PO4 backbone, decreases repelling and creates stronger interactions
103
DNA Mutagenesis
spontaneous mutagenesis
oxidative and chemical mutagenesis
radiation induced mutagenesis
104
DNA mutagenesis: spontaneous
deamination
depurination
105
DNA mutagenesis: spontaneous deamination
amino group converted to carbonyl
good to have thymine in DNA because uracil is detected as a problem (C--> U 100/day in mammalian cell)
slow reactions
106
DNA mutagenesis: spontaneous depurination
N glycosidic bond is hydrolyzed
10K purines lost a day
107
DNA mutagenesis: oxidative mutagenesis
hydroxylation of guanine
often in mitochondrial DNA because lots of oxidative phosphorylation takes place
108
DNA mutagenesis: chemical alkylation
methylation of guanine
109
DNA mutagenesis: radiation induced mutagenesis
UV light
ionizing radiation (X rays, gamma rays)
110
DNA mutagenesis: UV light
dimerization of pyrimidines
main mechanism for skin cancer
111
DNA mutagenesis: ionizing radiation
x rays
gamma rays
ring opens, strand breaks
112
accumulation of mutations leads to
aging and cancer
113
segments of DNA are arranged either within a ______ or between two _____
within a chromotome or between two DNA molecules giving offpsring novel DNA
114
DNA metabolism consists of
a set of tightly regulated processes that achieve these tasks
115
DNases
nucleases that cleave DNA (enzymes)
116
exonucleases
cleave a bond that removes a nucleotide from the end of DNA; can keep cleaving
117
endonucleases
cleaves a bond within a DNA sequence, between nucleotides, cleaves once and makes nick
118
3 fundamental rules of replication
semi conservative
begins at an origin and proceeds bidirectionally
synthesis of new DNA occurs in 5' to 3' direction and is semi-discontinuous
119
MESELSON-STAHL
cells grown on heavy medium with N15; found one band
cells on N14 medium divided once; one band on higher position
cells divided again in N14; two bands, one with N14DNA, one hybrid
semiconservative**
120
leading strand
made continuously as replication fork advances
121
lagging strand
made discontinuously away from replication fork in short pieces that are later joined together
okazaki fragments
122
DNA polymerases
synthesizes DNA, requires a primer
123
primer
short strand of NT complementary to template; provides required 3'OH to begin the DNA polymerase catalyzed reaction
124
substrates in strand synthesis
nucleoside triphosphates
125
how many phosphates removed during catalysis as one unit?
2; pyrophosphate
126
what is attached to growing DNA strand?
dNMP
127
what attacks the a-phosphate on incoming trinucleotide?
nucleophilic 3'OH on growing strand
128
Mg2+ purpose
makes 3'O on growing strand more powerful nucleophile by stabilizing it --> allows it to attack a-PO4
orients the components, making b and y PO4 better LG
129
how is Mg2+ in catalytic site of DNA polymerase?
interactions with aspartate groups
130
errors in E.Coli base pairing
1/10^9
131
ways to limit incorrect base pairing
base pair geometry at insertion site
errors during synthesis are corrected by 3'-5' exonuclease activity
132
base pair geometry
geometry limits incorrect base pairing bc doesnt fit in catalytic site
133
DNA polymerase has ___ and __
primary active site
secondary active site
134
primary active site in DNA polymerase
synthesis; polymerase
135
secondary active site in DNA polymerase
exonuclease activity
136
3' to 5' exonuclease activity
proofreads synthesis for mismatched repair
translocation of enzyme to next position is inhibited until the enzyme can remove the incorrect nucleotide just added
137
DNA polymerase I (E.Coli)
abundant, not ideal for replication
600 nt/min (slower than replication fork)
low processivity
for primer replacement
138
DNA polymerase III (E.Coli)
principle replication polymerase
139
DNA polymerases II, IV, V
DNA repair
140
DNA Polymerase I 5'to3' exonuclease
in addition to 3'to5'
activity has a distinct domain
works ahead of polymerase activity -- hydrolyzes nucleotides off chain in its path before adding new nucleotides with polymerase activity
nick translation
141
nick translation
DNA pol I
movement of strand break along with enzyme resynthesis
combined 5' to 3' exonuclease activity and polymerase activity
142
DNA polymerase III
10 types of subunits
two core polymerases made of a, e, theta subunits
clamp loader complex
143
clamp loader complex
DNA pol III
links core polymerases
144
core polymerases in DNA pol III
each interact with a dimer of beta subunits that increase the processivity of complex
145
sliding clamp
B subunits on DNA pol III form this; prevents dissociation
polymerase holds clamp, clamp holds DNA
146
processivitiy of DNA pol III
>500k bp
147
a subunit of core DNA polymerase on DNA Pol III
polymerization
148
e subunit of core DNA poly on DNA pol III
3'-5' proofreading exonuclease
149
theta subunit of DNA poly
stabilization of e subunit
150
DNA replication in E.COli
replisome
151
replisome
over 20 proteins required for replication in E.Coli
152
replisome proteins
helicase
topoisomerase
DNA binding proteins
primase
DNA ligase
153
helicase
cannot separate strands; continue separating strands, unwinds DNA with ATP
154
topoisomerase
relieves stress caused by unwinding; releases strain in front of forks by cutting DNA, relieves twist, then joins them again
155
DNA binding proteins
stabilizes separated strands
156
primase
enzyme that makes RNA primers 5-15 bases long; provides 3'OH
157
DNA ligase
seals nicks in backbone
158
Initiation of prokaryotes
begins at origin: oriC
highly conserved sequence elements
159
origin in prokaryotes
oriC
160
oriC
origin; 245 bp
5 repeats of a sequence that forms binding site for DnaA
A-T rich DUE
161
DnaA
initiator protein; first protein that binds origin
prokaryotes
162
DUE
DNA unwinding element; A-T rich
strands pop open and strands separate
with binding sites for DnaA and other proteins
163
prepriming complex
goal after opening helix in prokaryotes
164
DnaA
recognizes ori sequence, opens duplex at specific sites in origin
p
165
DnaB
helicase; unwinds DNA
166
DnaG
primase; synthesizes RNA primers
167
SSBP
binds single stranded DNA
168
DNA gyrase
DNA topoisomerase II
relieves torsional strain
169
DnaA proteins binding at oriC
ATPases
8 DnaAs bind
DNA wraps around 8 protein complex forming a positive supercoil
strain leads to denaturation of DNA in nearby DNA unwinding element, overcomes base stacking
170
Elongation in prokaryotes
DNA polymerase III adds beta clamp at primer and core polymerase binds to b clamp
adds deoxynucleotides in 5' to 3' direction
lagging strand, leading strand synthesis -- SAME DNA POLY
171
lagging strand synthesis
okazaki fragments
DNA polIII adds NT in short segments, ends when hits previous primer
172
connecting okazaki fragments
core poly of DNA polIII dissociates from one b clamp, binds to a new one
DNA pol I
DNA ligase
173
DNA pol I with okazaki fragments
associates with b clamp, and nick translates
5'to3' exonuclease to remove primer
polymerase to replace primer with DNA (and on leading strand)
174
DNA ligase
makes bond between a 3'OH and a 5'PO4
175
DNA ligase mechanism
removes AMP from ATP or NAD+
attach AMP on lysine on ligase (itself)
attaches AMP onto 5'PO4 on strand
3'OH attacks 5'PO4, AMP is displaced
176
Termination in prokaryotes
replication forks meet within a region of Ter sequences
177
Ter sequences
Ter sites found near each other but in groups oriented in opposite directions near tus proteins
create a site that replication forks cannot leave
178
Tus proteins
terminus utilization sequence
binds ter sites (not all are bound by tus)
catches the fastest fork and waits for catch up
179
yeast have X defined origins
400
180
why do eukaryotes have more origins?
more chromosomes
181
origin recognition complex (ORC)
in eukaryotes; loads a helicase onto DNA
hexamer of mini-chromosome maintenance proteins
similar to DnaA
182
eukaryotic replication
slower
50 nt/sec (250 in E.Coli)
compensated by origins every 30-300 kb
183
termination occurs in eukaryotes when
replication forks converge
184
DNA polymerase alpha primase complex
in eukaryotic replication
all removed after replication is over
makes primers with RNA, then 10nt DNA
separates subunits for primase and polymerase activities
no 3'-5' proofreading
185
DNA polymerase delta
synthesizes lagging strand
3'-5' proofreading
binds to PCNA protein
186
PCNA
proliferating cell nuclear antigen
acts like a beta clamp to increase processivity in DNA polymerase delta
187
DNA polymerase e
synthesizes leading strand
3' to 5' proofreading
binds to PCNA to increase processivity
188
messenger RNA
encodes the aa sequences of all polypeptides found in cell
189
transfer RNA
match specific aa to triplet codons of mRNA during protein synthesis
190
ribosomal RNA
constituents of ribosomes along with proteins
191
ribonucleic acids
act as genomic material in viruses
192
transcription
ribonucleic acids are synthesized in cells using DNA as a template
tightly regulated in order to control the concentration of RNA and save energy
193
many RNA molecules can
fold into structures with functions
194
most RNAs are processed after synthesis
elimination of introns, joining of exons
poly-adenylation
capping
195
Transcription in E.Coli: RNA polymerase
catalyzes synthesis in 5'to3' direction
doesnt need primer
first nucleotide retains all 3 phosphates on 5' end
NTPs are used to add NMPs to the 3' end of growing strand
196
RNA polymerase in E.Coli
binds promoter
opens up and unwinds DNA (unwound duplex forms a bubble 17bp long)
topoisomerase releases strain
growing end of new RNA temporarily base pairs with DNA template for 8 bp
197
RNA polymerase is a multisubunit
holoenzyme
5 core subunits a2bb'w and sigma as a sixth
no 3'to5' exonuclease; high error rate, does proofread though
198
sigma subunit on RNA polymerase
directs enzyme to the promoter
diff subunits bind diff promoters
199
promoters in E.Coli
sequences in DNA where RNA polymerase binds with sigma subunit
200
two promoter consensus sequences in E.Coli
TATA sequences
-10 (TATAAT) and -35 (TTGACA) for sigma subunit binding
201
UP element
AT rich upstream promoter element
between -40 and -60 binds the alpha subunit
promote strand separation
202
AT rich upstream promoter elements promote strand separation
sequences govern efficacy of RNA poly binding, affecting gene expression binding
not every promoter has this; differences impact how good the polymerase binds
203
sigma 70
housekeeping, many molecules present at a time
204
sigma 32
heat shock genes
205
holoenzyme ratio
how much RNA polymerase binds to that sigma subunit
206
DNA footprinting
a way to find the binding site on DNA for a DNA binding protein
DNA bound by a protein will be protected from chemical cleavage at its binding site
207
DNA footprinting steps
1. isolate DNA fragment with binding site
2. radiolabel DNA and split sample into two tubes
3. add binding protein to DNA in one tube
4. protein will bind to sequence
5. treat both tubes with chemical or enzymatic agent to cleave DNA into many pieces
6. separate fragments by gel electrophoresis
if protein binds, multiple bands will be missing in the lane for that sample --> where RNA poly bound to DNA
208
transcription initiation: RNA polymerase
uses the sigma subunit to separate the DNA strands without helicase or primers
209
closed complex
RNA polymerase binds to promoter with sigma subunit, creating closed complex because DNA is not unwound
210
open complex
region of DNA from -10 to 2 is separated
211
sigma subunit holds the RNA to the promoter and
must dissociate to allow elongation
212
transcription elongation
not as efficient as DNA polymerase
RNA polymerase adds NMPs; can stall or backtrack
213
RNA polymerase proofreading
moves backwards which separates the 3' end of RNA
RNA probably cleaved in catalytic site of RNA polymerase and resynthesized
GreA and GreB
214
GreA and GreB
elongation factor binding to RNA polymerase increases efficiency of RNA poly proofreading
215
two types of termination in prokaryotes
rho-independent termination in E.Coli
rho-dependent termination in E.coli
216
Rho-independent termination in E.Coli
intrinsic
self complementary regions in mRNA transcript form a hairpin 15-20 nt before 3' end
makes RNA poly pause
RNA dissociates from DNA due to poly-U region
217
why does the RNA dissociate from DNA at the poly-U region?
A-U binding is weak
218
rho-dependent termination in E.Coli
rho-helicase binds to Rut site on RNA
proceeds 5'to3' until it reaches paused RNA poly
uses helicase activity to release RNA from DNA
219
rut site
rho utilization element on mRNA
220
regulation of transcription
regulate affinity of RNA poly for a promoter
alter promoter sequence
activator proteins make RNA poly more likely to bind
repressor proteins make RNA poly less likely to bind
221
Eukaryotic RNA polymerases
RNA pol I
RNA pol II
RNA pol III
RNA pol IV
222
Eukaryotic RNA pol I
synthesizes pre-reibosomal RNA
precursor for rRNAs
223
Eukaryotic RNA pol II
responsible for synthesis of mRNA
very fast (500-1k nt/sec)
can recognize thousands of promoters
224
Eukaryotic RNA pol III
makes tRNAs and some small RNA products
225
Eukaryotic RNA polymerase IV
some plants, synthesizes siRNA
226
Eukaryotic mRNA transcription: binding at promoter
relies on protein-protein contacts, with highly conserved tfs
227
RNA polymerase II in eukaryotic transcription
large complex of 12 subunits
largest subunit has a carboxy-terminal domain (CTD)
228
CTD
contains highly conserved repeats which are phosphorylated and dephosphorylated to control transcription
eukaryotic
229
Eukaryotic initiation of transcription
TATA binding protein (TBP) binds to the promoter
230
TATA binding protein is similar to
sigma subunit in prokaryotes
231
TFIIH functions
helicase activity
| kinase activity
232
TFIIH helicase activity
unwinds DNA at promoter
233
TFIIH kinase activity
other kinases phosphorylate the CTD of RNA pol II
changes conformation and enables RNA pol II to transcribe
234
Elongation factors in Eukaryotes
bound to RNA pol II
enhance processivitiy
allow correction of incorrect base addition
coordinate post-transcriptional modification
235
RNA pol II is dephosphorylated for termination
once the RNA is out of RNA pol II, protein complex that cleaves bind to the CTD, cleave RNA and add a poly-A-tail before polymerase finishes making mRNA
doesn't matter where termination occurs
236
Rna pol phosophorylated =
transcription
237
Rna pol dephos
termination
238
5' cap
7-methylguanosine
links to the 5' end of mRNA
239
7'methylguanosine links to 5' end of mRNA via
5',5'-triphosphate link
may include additional methylations at 2'OH group sof next two nucleotides
240
functions of 5'cap
protects RNA from exonucleases
forms a binding side for ribosome
added during transcription by complex of 4 enzymes
241
how is 5' cap added to mRNA during transcription?
4 enzymes attached to CTD of RNA pol II
as mRNA exits polymerase, cap is put on
242
introns
segments not part of DNA
50-20k nt
243
classes of introns
group 1 and 2 (selfsplicing)
spliceosomal introns
tRNA introns
244
group 1 and 2 introns
self-splicing
ribozymes that cleave themselves or another RNA
3D structure integral to function
MM kinetics
no additional proteins or ATP
245
genomes of group 1 and 2 introns
nuclear, mito, chloro
246
group 1 and 2 introns differ in
splicing mechanism
247
spliceosomal introns
spliced by spliceosomes
most common introns
frequent in protein-coding regions of eukaryotic genomes
248
spliceosomes
complexes with RNA and proteins
associates with the CTD of RNA pol II, splices as RNA is made
249
tRNA introns
spliced by protein based enzymes
primary transcript cleaved by endonuclease
exons joined by ATP ligase
250
poly A tail
RNA pol II synthesizes the mRAN past cleavage site
protein complex binds mRNA at cleavage site
endonuclease recognizes site, cleaves RNA 10-30nt downstream from AAUAA
added by polyadenylate polymerase
subject to exonuclease activity
251
polyadenylate polymerase
synthesizes 80-250 nt of adenine without a template onto 3'OH at end of mRNA with AAUAA site
252
how to get diff mature transcripts from gene?
RNA processing
location determines expression
253
average RNA half life
3 hours
10 turnovers/cell generation
254
bacterial cells half lives
shorter (1.5 min)
255
ribonucleases
degrade mRNA
256
endoribonuclease
clips RNA into segments
257
exoribonuclease
breaks down fragments generated by endoribonuclease or degrades whole RNA strand at either end
5' cap must be removed in eukaryotes via decapping enzymes before 5'-3' exoribonuclease activity
258
why is protein synthesis controlled?
it is energy demanding
uses 90% of chemical energy in the cell
259
how many proteins does protein synthesis require?
>300 biomolecules in eukaryotes
260
proteins are synthesized within
ribosomes attached to cytosolic face of ER
261
amino acids are activated for synthesis by
attachment to tRNA
262
how are AA attached to tRNA?
aminoacyl-tRNA synthetases
263
tRNA acts as an ____ to translate mRNA into protein
adapter
264
first codon
establishes reading frame
265
degenerate code
61 out of 64 codons code for 20 amino acids
266
initiation codon
AUG
267
termination codons
3 out of 64; UAA UGA UAG
268
genetic code in mitochondria
contain DNA to make a few proteins but with a slightly different code
encode their own tRNAs and use 22 instead of 32
269
silent mutations
diff NT in DNA or mRNA but same AA in protein
270
conservative substitution
in some cases where mutation in the first base of a codon encodes an AA with similar characteristics
GUU -- valine
AUU -- isoleucine
CUU -- leucine
271
how does the codon in mRNA base pair with the anticodon on tRNA?
hydrogen bonding, antiparallel
272
wobble base pair
third base of a codon of mRNA can form non-canonical base pairs with complement in tRNA
G can bind U
273
inosinate (I)
can bind A C U non canoncially
274
how many tRNAs properly translate 61 codons?
32
275
are there tRNAs for stop codons?
no
276
5 stages of p synthesis
activation of AA
initiation of translation
elongation
termination and ribosome recycling
folding and post translational processing
277
5 stages of p synthesis:
activation of AA
by binding to tRNA
end result: tRNA is aminoacylated
278
5 stages of p synthesis:
initiation of translation
mRNA and first aminoacylated tRNA bind to ribosome
279
5 stages of p synthesis:
elongation
cycles of aminoacyl-tRNA binding and peptide bond formation until stop codon
280
5 stages of p synthesis:
termination and ribosome recycling
stop codon enters ribosome, protein leaves ribosome, ribosome and mRNA recycled
281
5 stages of p synthesis:
folding and post translational processing
catalyzed by a variety of enzymes
282
tRNA features
73-93 nt in both bact and euk
2D: cloverleaf
3D: twisted L
G at 5' end
CCA at 3' end
283
aminoacyl-tRNA synthetase
binds a specific amino acid to its matching tRNA
20 in each cell, for each AA
specific for AA and tRNA
284
second genetic code
matching each AA with the correct tRNA
285
how is the AA linked to the tRNA?
through the carboxyl group of AA to the 3' O of the last nucleotide on the 3' end of tRNA
286
synthesis of aminoacylated tRNAs steps
1. creation of aminoacyl intermediate
2. transfer of aminoacyl to tRNA
all in catalytic site of aminoacyl tRNA synthetase
287
synthesis of aminoacylated tRNAs steps:
creation of intermediate
aminoacyl-tRNA synthetases esterify 20 AA to corresponding tRNAs
carboxyl of AA attacks alpha PO4 of ATP, creating intermediate
pyrophosphate is cleaved and becomes product
288
what drives the creation of aminoacyl intermediate?
two phosphoanhydride bond cleavages
breaking bond between AMP and pyrophosphate
AND
energy from breaking bond from two inorganic phosphates
289
synthesis of aminoacylated tRNAs steps:
transfer of aminoacyl to tRNA
2'OH or 3'OH of tRNA attacks the carbonyl carbon of the amino acyladenylate intermediate
creates ester bond between AA to the 3'O of the tRNA
290
what if the 2'OH attacks the carbonyl carbon of the aminoacyladenylate intermediate?
3'O attacks the carbon to transfer the AA
291
ribosome makes up ___ weight in bacteria
25%
292
ribosome structure
65% rRNA, 35% protein
2 subunits bound together
293
rRNA
forms the core of ribosome, does catalysis of peptide formation
294
does protein catalyze peptide bond formation?
NO, rRNA does
295
ribosome subunits bacteria
50s and 30s = 70s
296
ribosome subunits eukaryotes
60s and 40s = 80s
297
why is there a large subunit of ribosomes?
heavier; bigger, sediments down in centrifugation
298
rRNA secondary structure
three single stranded rRNAs of E.Coli have specific 3D structure with intrachain base pairs
shape of rRNAs is highly conserved (bacteria, archea, eukaryotes)
299
first codon is
AUG
300
all organisms have ____ tRNAs for Met
2
1 for initiating tRNA and one for an internal tRNA
301
bacteria Met
N-formylmethionine; fMet
initiation tRNA: tRNAfmet
interior met: normal tRNAmet
302
eukaruotes Met
protein begins with Met BUT a special initiating tRNA is used
303
bacteria initiation of translation requires
small 30S ribosomal subunit
mRNA
fmet-tRNA (initiating tRNA)
initiation factors
GTP
large 50S ribosomal subunit (for end of initiation)
304
initiating factors required for bacterial translation
IF1
IF2
IF3
305
prokaryotic initiation
can start during transcription
306
prokaryotic initiation:
| steps
30S subunit bound by IF3, IF1
mRNA binds small subunit
initiating 5'AUG codon is guided to correct position in robosome via shine-dalgarno sequence
IF2 helps tRNA fmet bind in P site
IF3 departs
large subunit binds
initiation complex is completed
IF2 hydrolyzes GTP to GDP
IF1 and IF2 dissociate
307
what directs 5'AUG initiating codon to correct ribosomal position?
shine-dalgarno sequence
308
shine-dalgarno sequence
region in mRNA that is complementary to a sequence in rRNA
16s rRNA
309
IF3
keeps 30s subunit and 50s apart
310
IF1
blocks the A site to prevent tRNA binding accidentally
311
IF2
bound to GTP, binds small subunit at P site
helps formulmethionine-tRNAfmet bind in the P site to initiating AUG
312
initiating complex in prokaryotes
all subunits bound
313
at the end of initiation
complete ribosome with mRNA bound and the initiating tRNA in the p site
314
initiation of translation in eukaryotes
elF2 ginds met-tRNAmet and brings it to P site of small (40S) subunit
elF4F binds mRNA and helps it bind to small subunit
small subunit moves along mRNA until AUG is in the P site
then binds initiating methionine tRNA
initiating factors leave, 60s subunit binds
315
elF1A
IF1
blocks A site to prevent tRNA binding accidentally
316
elF1
IF3
keeps small and large subunit separate
317
elF4F
orients mRNA by binding of 5'cap to rRNA
| SD sequence similarity
318
elongation in prokaryotes
next aminoacyl tRNA binds to EF-Tu
one AAtRNA in A site, one in P site
N-formylmethionine or growing peptide is transfered from tRNA in P site to AA in A site
peptide bond formation catalyzed by 23S rRNA
deacetylated tRNA in P site
ribosome translocation
319
EF-Tu
elongation factor Tu
docks amino-acyl tRNA EF Tu GTP at A site of 70S initiation complex
hydrolyzes GTP, leaves
recycled
320
how does EF-Tu recycle
interactions with EF-Ts by loss of GDP and binding of GTP
321
23S rRNA
catalyzes peptide bond formation in prokaryotes
322
elongation in prokaryotes:
ribosome translocation
ribosome moves one codon toward the 3' end of mRNA
uses energy from hydrolysis of GTP on EF-G
uncharged tRNA now in E site and can leave ribosome
growing peptide chain on tRNA in p site
A site open for new AA tRNA
323
EF-G
binds at A site, hydrolyzes GTP, leaves, recycles
324
eukaryotic elongation factors
eEF1alpha
eEF1by
eEF2
325
eEF1alpha
EF-Tu
binds to aminoacyl tRNA and docks at the A site of large subunit; hydrolyzes GTP and is recycled
326
eEF1by
EF-Ts
| interacts with eEF1alpha and recycles it
327
eEF2
EF-G
hydrolyzes GTP in A site for translocation, leaves
328
termination in prokaryotes
elongation continues until a stop codon reaches the A site
UAA, UAG, or UGA on mRNA triggers action of termination factors
329
Termination factors prokaryotes
RF1/RF2
| RF3
330
RF1/RF2
hydrolyzes terminal peptide-tRNA bond to release peptide from tRNA
each one binds different stop codons
331
RF3
releases RF1 or RF2 from the ribosome
332
ribosome recycling
binding of ribosomal recycling factor (RRF) and EF-G and hydrolysis of GTP
333
binding of ribosomal recycling factor (RRF) and EF-G and hydrolysis of GTP causes...
exit of tRNAs
dissociation of large subunit from small subunit
release of mRNA
IF3 binds again to small subunit to prevent reassociation of ribosomal subunits
334
termination in Eukaryotes
eRF1
| ABCE1
335
eRF1
RF1/RF2
recognizes all 3 stop codons and releases peptide from tRNA via hydrolysis
336
ABCE1
ATP-binding cassette subfamily E member 1
interacts with eRF1 and separates the subunits of ribosome
requires ATP hydrolysis, part of recycling
337
protein synthesis features
large energy cost
can be rapid when accomplished on polysome
in bacteria, tightly coupled to transcription
338
large energy cost of protein synthesis
1 ATP to put AA on tRNA with aminoacyl-tRNA synthetases
2 GTP/amino acid (one for EF-G ribosome translocation, one for EF-Tu for AA-tRNA bind to A site)
1 GTP for initiation (IF2 hydrolyzes after helping initiating tRNA bind in P site)
1 GTP for termination (recycling of EF-G after translocation)
339
polysome
clusters of ribosomes
340
posttranslational modifications
removal of formyl group on first residue or removal of Met and other N term residues
acetylation of N term residue
removal of signal sequences or other regions
attach. carbs
modify AAs with additional COO-, CH3, or OH
addition of isoprenyl groups or other lipids (anchoring in membranes)
adding cofactors
forming disulfide links
proteolysis of proproteins
341
signal sequences
direct proteins from site of synthesis; at or near N terminus
342
as peptide emerges from the ribosome, signal sequence is bound by a
signal recognition particle (SRP)
made of RNA and proteins
343
SRP binds GTP and
inhibits translation of protein
344
SRP / peptide / ribosome / RNA complex goes to ER
SRP helps bind outside of the ER, dissociates with GTP hydrolysis
protein translation finishes
stays in the ER membrane or translated through
signal peptide is cleaved, modification takes place
345
transport vesicles
take proteins from ER to golgi if protein is destined for another organelle or for secretion
346
proteins for mito and chloroplasts bind ________ in cytosol and are delivered to ___________ on the organelle
chaperone proteins; receptors on the exterior of the organelle
347
proteins destined for the nucleus have
a nuclear localization sequence (NLS)
348
NLS
not cleaved after protein is targeted because the nuclear envelope can break down during mitosis and proteins will need to reenter the nucleus
349
how does NLS go to nucleus?
binds importin alpha and beta and RanGTPase
complex docks at a nuclear pore and is imported
350
half life of metabolism regulatory proteins that respond to rapidly changing needs
short lived
351
biological functions of lipids
energy storage
insulation
water repellant
buoyancy control and acoustics in marine animals
membrane structure
signaling molecules
pigments
antioxidants
352
biological functions of lipids:
energy storage
composed of. reduced compounds for available energy
hydrophobicity allows lots of packing bc of no water
353
biological functions of lipids:
insulation
low thermal conductivity
high heat capacity
mechanical protection
354
biological functions of lipids:
water repellent
hydrophobicity keeps surface of organism dry
no wetting
no evaporation
355
biological functions of lipids:
signaling molecules
paracrine (local)
steroid (body wide)
growth factors
Vitamins A and D (hormone precursors)
356
biological functions of lipids:
pigments
tomatoes, carrots, pumpkins, birds, leaves
357
classification of lipids
do not contain fatty acids
do contain fatty acids
358
lipids that do contain fatty acids
storage lipids (neutral)
membrane lipids (polar)
359
fatty acids
carboxyllic acids with hydrocarbon chains with 4-36 carbons
most 12-24
even number of carbons
360
most natural fatty acids are
unbranched
361
levels of saturation in fatty acids: mono
one c=c bond
362
conformation of fatty acids
saturated chains tend to adopt extended conformations
double bonds in natural unsaturated fatty acids are in cis conformation which kinks the chain
363
essential nutrients
things we cannot synthesize
omega 3
omega 6 fatty acids
364
as the chain length increases, solubility
decreases because there are more hydrophobic interactions
365
as the chain length increases, the melting point
increases because there are more interactions which require more energy to melt
366
as the number of double bonds increases, the melting point
decreases because they are packed less tightly and have less interactions
367
saturated double bonds pack in a. _______ and have a _____ Tm
more orderly way and have a higher Tm
368
unsaturated fatty acids with cis c=c pack ______orderly due to kink
less;
less thermal energy to disrupt disordered packing, lower Tm
369
how do trans fatty acids form?
partial hydrogenation of unsaturated fatty acids
done to increase stability at high temperature of oils used in cooking or longer shelf life
370
trans fatty acids
allows a fatty acid to adapt an extended conformation like a saturated fatty acid
371
triacylglycerols
storage lipid
three fatty acids form ester linkages with three hydroxyl groups of glycerol
majority of FA in biological systems in this form
primary storage form of lipids in organisms
372
solid triacylglycerols
fats
373
liquid triacylglycerols
oils
374
properties of triacylglycerols
less soluble in water than fatty acids due to lack of charged carboxyl group
less dense than water, fats and oils float
375
advantage of fats over polysaccharides
carry more energy per carbon because they are more reduced sugars
carry less water per gram because are non polar and pack tightly
376
short term energy needs
glucose and glycogen, storage for carbs, quick delivery
377
long term energy needs
fats, good storage, slow delivery
378
waxes
esters of long chain sat or unsat fatty acids with long chain alcohols
379
wax properties
insoluble, high melting points
380
functions of waxes
protection and pliability for hair and skin in verts
waterproofing of feathers
protection from evaporation in plants
used by people in lotions and ointments etc
381
structural lipids in membranes
polar head groups and nonpolar tails (typically fatty acids)
382
diversification of membrane lipids comes from
different backbone
different fatty acids
addition of head groups
383
what determines the surface properties of membranes?
properties of. head groups
384
glycerophospholipids
type of lipid in membranes
primary constituents of cell membranes
two FA form ester linkages with first and second hydroxyl groups of L-glycerol-3-phosphate
head group on phosphate group
unsat FA on C2
385
phosphatidylcholine
most common glycerophospholipid
major component of most cell membranes (euk)
not in prokaryotes, cant be synthesized
386
sphingolipids
second type of lipids in membranes
backbone is sphingosine (long chain amino alcohol)
FA joined to sphingosine via amide linkage
head group on sphingosine via glycosidic or phosphodiester linkage
387
where are sugar-containing glycosphingolipids often found?
outer face of plasma membrane
388
blood groups are determined by
type of sugars located on the head groups in glycosphingolipids
389
the structure of the sugar in head group of glycosphingolipid is determined by expression of specific
glycosyltransferases
390
no glycosyltransferases?
o antigen
391
glycosyltransferase for N-acetyl galactosamine?
A antigen
392
glycosyltransferase for galactose?
b antigen
393
sterols
cholesterol = most common
steroid nucleus of 4 rings, almost planar
hydroxyl group/polar head in A ring
various nonpolar side chains
394
physiological role of sterols (cholesterol)
present in the membranes of. most eukaryotic cells
modulate fluidity and permeability
thicken membranes
395
bacteria lack
sterols
396
how do mammals obtain cholesterol?
from food or synthesize in liver
397
cholesterol is transported to tissues in
lipoproteins
398
lipoproteins
transports choelsterol
combination of lipids and proteins
categorized by density
399
small particles have
high density
400
large particles have
low density
401
low-density lipoproteins
tend to deposit and clog arteries
402
physiological role of sterols (hormones)
hormones are derivatives of sterols
