BB450 exam 2 Flashcards
proteases
enzymes that cleave peptide bonds
catalyze addition of water
chymotrypsin is a ___ protease
serine
catalytic triad
serine H - histidine - aspartic acid
catalysis of serine proteases
- binding of substrate
- conformational change –> alkoxide ion
- attack of alkoxide ion on carbonyl carbon of peptide bond –> unstable intermediate
- stabilization of intermediate –> breaking peptide bond (one side attached to serine, other side released)
- water enters active site
- activation of water by removal of proton from histidine
- attach of hydroxyl on carbonyl carbon attached to serine
- release of second peptide and regeneration of original active site
alkoxide ion formation
binding of substrate brings histidine closer to hydroxyl group of serine and extracting protein
oxyanion hole
stabilizes tetrahedreal intermediate that arises twice during catalysis, keeps intermediate from reacting with enzymes (adjacent to active site/catalytic triad)
S1 pocket
where substrate binds and catalysis occurs (adjacent to active site/catalytic triad)
determines serine protease’s specificity
S1 pocket of chymotrypsin
hydrophobic and relatively large (to bind to phenylalanine)
S1 pocket of trypsin
negatively charged to bind to lysine or arginine
chymotrypsin cuts at
adjacent to phenylalanine (and other hydrophobic aa)
aspartyl proteases
use 2 aspartic acid side chains to hold water in place; use ion to act as nucleophile to attach peptide bond
metalloproteases
use metal ion to hold water in place so can be ionized to act as nucleophile to attach peptide bond
Carbonic Anhydrase
enzyme that catalyzes joining of CO2 and water to form carbonic anhydrase
carbonic anhydrase process
zinc ion held by 3 histidines in active site bind water molecule
loss of proton my water –> catalysis
carbonic anhydrase max effective at
high pH - protons easily removed
limiting step in carbonic anhydrase
abstraction of proton from water
buffers and/or bases help facilitate this
restriction enzymes/endonucleases
bacterial enzymes that can cleave DNA by breaking phosphodiester bond between adjacent nucleotides
restriction enzymes are defense against ___
viruses
___ prevents restriction enzyme from cutting DNA
methyl group (mathylase puts methyl group on nucleotide) cellular, methylated DNA remains uncut, invading viral DNA (not methylated) gets cut and destroyed
Myosins
translate ATP energy into movement
ATCase catalyzes
1st reaction in CTP synthesis (pyrimidine synthesis)
*does not synthesize CTP
substrate of ATCase
aspartate - substrate at active site
indicates cell’s readiness to divide
hold in R state (active)
CTP inhibits
ATCase - feedback inhibition (mediated allosterically)
CTP locks it in T state (can’t flip to R anymore)
____ activates ATCase
ATP and aspartate
ATP and ATCase
activates
ATP is a purine, indicates high energy of cell–> cell is ready to divide
hold is R state (active)
structure of ATCase
12 subunits: 6 catalytic, 6 regulatory
ATP and CTP bound at regulatory subunit
aspartate bound at catalytic substrate at active site
PALA
binds to active site of ATCase covalently
suicide inhibitor
locks enzyme in R state, blocks access to active site
acts like aspartate (proves aspartate is causing to be in R state)
concerted model
ATCase can flip between R and T state all by itself, allosteric effects lock it in state
no cause/effect sequential model like hemoglobin
Protein Kinase A
attaches phosphates to serine or threonine
controlled by allosteric means
2 regulatory, 2 catalytic subunits: R2C2
control whether enzymes are active or inactive
when catalytic subunits bind to regulatory subunits of Protein Kinase A
cannot catalyze reactions
when cAMP binds to regulatory subunits of Protein Kinase A
catalytic subunits released and active
phosphatases
remove phosphates
cAMP
signaling molecule, activated Protein Kinase A
Protein Kinase A activates enzymes involved in…
breakdown of glycogen and synthesis of glucose
in liver
epinephrine–>cAMP–>protein kinase A–>phosphorylates enzymes–>activates production of glucose
phosphodiesterase
breaks down cAMP
stopping breakdown of glycogen and reducing blood glucose to normal
caffeine…
inhibitor of phosphodiesterase, favors high blood glucose levels
zymogens
enzymes that are synthesized in an inactive form
activation requires covalent modification, usually proteolytic cleavage
digestive enzymes
trypsin, chymotrypsin, elastase, carboxypeptidase
proteolytic enzymes
break down proteins, needed for digestion
take ___ to activate a protease
another protease
pancreatitis
proteolytic enzymes attack pancreas (where they’re made) overly active, get activated closer to pancreas
___ is primary activator or proteolytic enzymes
trypsin
chymotrypsinogen –> chymotrypsin
requires trypsin
makes initial cleave between aa 15 and 16 (disulfide bonds keep them from coming completely apart)
becomes pi-chymotrypsin, cleaves itself to remove 2 dipeptides–> full chymotrypsin activity (3 polypeptide pieces held together by disulfide bonds)
pi-chymotrypsin
intermediately active form
after trypsin makes initial cleave between aa 15 and 16
alpha one antitrypsin
protease inhibitor that stops elastase in lungs from getting too active
(plugs up active site of trypsin, works better on elastase but already got name)
elastase
breaks down proteins we breath in
is elastase is too active –>
emphysema
smokers and alpha one antitrypsin
oxidize methionine in alpha one antitrypsin
prevents it from binding to elastase
elastase too active –> emphysema
blood clotting general
2 pathways possible
blood clot is self assembly of fibrin
blood clotting pathway
prothrombin bind to calcium and held at wound site
prothrombin –> thrombin
thrombin converts fibrinogen –> fibrin
fibrin polymer hardens by glutaminase
fibrinogen
alpha, beta, and gamma chains
dimer
alpha has A on end, beta has B on end
fibrinogen –> fibrin
thrombin clips A and B portions of alpha and beta of fibrinogen, polymerization
alpha left over fits into gamma
beta left over fits into hole on beta structure
transglutaminase (glutaminase)
hardens fibrin polymer
combines side chains of glutamine and lysine with covalent bond
covalent bond gives strength to clot
binding of prothrombin to calcium…
anchors prothrombrin in phospholipid membranes derived from blood platelets after injury
prothrombin converted to thrombin at this site
enable prothrombin to bind calcium…
glutamate resides must be carboxylated
this reaction catalyzed by enzyme that uses vitamin K as cofactor
block action of vitamin K…
anticlotter
how blood thinners works (Coumadin, warfarin)
competitive inhibitor: compete with vit. K for active site
plasmin
removal of blood clots
synthesized by plasminogen
t-PA
tissue type plasminogen activator
plasminogen –> plasmin
effective in inhibiting cascade to dissolve unwanted clot from stroke or heart attack
serine protease
glucose is an ___
aldohexose
most abundant sugar
glyceraldehyde
aldo-triose
simplest saccharide we call carb
dihydroxyacetone
keto-triose
simplest saccharide we call carb
D
2nd to last hydroxyl on right
most biological sugars
L
second to last hydroxyl on left
enantiomers
non superimposable mirror images
diasteriomers
differ in sterioisomeric configuration, not mirror images
epimers and anomers
epimer
differ in configuration of 1 carbon
anomer
differ in configuration of anomeric carbon
isomer
same formula, different structure
enantiomers and diasteriomers (epimers and anomers)
furanoses
5 carbon rings
pyranoses
6 carbon rings
ring structure arise from…
formation of hemiacetals or hemiketals
cyclization creates__
a new asymmetric carbon = anomeric carbon
carbon that was aldehyde or ketone
anomeric carbon can be ___
alpha (down) or beta (up) configuration
if hydroxyl group on anomeric carbon is unaltered
ring and linear forms can reversibly form
if hydroxyl group of anomeric carbon is altered (my methylation)
linear structure cannot form and flipping cannot occur
glycoside
altering hydroxyl group on anomeric carbon
commonly created during formation of disaccharides and longer carbs
form glycosidic bonds
aldehydes
very reactive, readily oxidized
ketones not…
not readily oxidized
chair and boat
different conformations
chair favored because less steric hindrance
sucrose
glucose in alpha + fructose in beta
non reducing
table sugar
lactose
glucose + galactose
B- 1,4 linkage
reducing sugar
maltose
glucose + glucose
alpha- 1,4 linkage
reducing sugar
reducing sugar
has free anomeric hydroxyl
lactose and glucose
in presence of copper 2+, copper will be reduced –> color change
oxidize aldehyde –>
acid
ribose linear structure
right, right, right,
deoxyribose linear structure
right, right, H
glucose linear structure
right, right, left, right
mannose linear structure
right, right, left, left
galactose linear structure
right, left, left, right
fructose linear structure
right, right, left, ketone
right is…
down in ring
left is…
up in ring
glycogen
energy stores for animals
polymer of glucose (a-1,4) with branches (a-1,6) ~every 10 residues
branches have branches
most in liver and muscles
cellulose
structural support in plants
only glucose
animals don’t make
b-1,4 linkages
starch
energy stores in plant, mixture of polysaccharides (amylose and amylopectin)
chitin
exoskeleton of insects
amylose
a-1,4 linkages of glucose, plants
one component of starch
homopolymers
contain only 1 sugar residue
glycogen, cellulose, amylose, amylopectin, chitin
amylopectin
a-1,4 links with a-1,6 branches ~every 30-50 residues
one component of starch
amylopectin/starch not as branched as glycogen…
less ends –> slower breakdown into glucose than glycogen because plants need less energy
cellulase
enzyme that breaks down b-1,4 linkages of cellulose
most animals don’t have it/cant digest cellulose
ruminants (cow, sheep) and ungulates have bacterium that make enzyme is specialized stomach
glycosaminoglycans
polysaccharides that contain amine group…
polyanionic –> chemical properties
proteoglycans
complexes or proteins and glycosaminoglycans that form feathery structures (repel each other bc negative charges)
slippery ex: snot and synovial fluid
glycoproteins
protein linked to oligosaccharide
N linkages
through N of asparagine
have common core of 5
made in Golgi or ER
O linkages
across serine or threonine oxygen
made only in Golgi
proteins from ER–> Golgi
additional modifications
targeting to cell membrane, release from cell, lysosome depending on patter on oligosaccharide surface
dolichol phosphate
embedded in ER membrane
N glycoprotein core attaches here
oligosaccharides to be linked with proteins
built on dolichol phosphate on outer portion of ER
then flip inside for attachment
hemagluttanin
protein on surface of flu viruses
bind specific carb. residues of surface of glycoproteins of blood cells
to exit cell, virus…
must cleave the sialic acid off w/ neuraminidase enzyme on virus surface
anti flu drugs
inhibit action on neuraminidase
cant exit cell after it divides
ex: Tamiflu
Haworth structure
ring form
Fischer projection
straight chain form
first messengers
epinephrine/adrenaline
insulin
epidermal growth factor (EGF)
second messengers
cAMP cGMP Calcium (3rd) DAG PIP3
response of signal include
enzyme activity, gene expression, cell division
7TM Receptors
cross cell membrane 7 times
N terminus sticks out, C terminus in cytosol
ex: beta-adrenergic receptor, rhodopsin
G protein
protein partner of 7TM receptor
on inside of cell that helps transmit signal
3 subunits: alpha, beta, gamma
G proteins can bind…
guanine nucleotides: GTP or GDP
beta-adrenergic receptor
- epinephrine binds to receptor
- G protein interacts with carboxyl tail of receptor and senses change
- alpha subunits binds GTP, releases GDP
- alpha subunit separate from b and gamma
- a subunit moves to adenylate cyclase
- conversion of ATP to cAMP
- cAMP interacts with protein kinase A–>phosphorylation–>produce glucose
alpha subunit of G protein bound to ___ when no signal present
GDP
when signal is present G protein ..
bind GTP and alpha subunits separates from beta and gamma
alpha subunits moves to..
adenylate cyclase
adenylate cyclase
membrane bound enzyme
catalyzes conversion of ATP to cAMP
turn off beta-adrenergic system..
GTP converted back to GDP
dissociation of epinephrine from receptor
receptor kinase to phosphorylate the carboxyl tail
breakdown of cAMP by phosphodiesterase
receptor kinase…
phosphorylates carboxyl tail of beta-adrenergic receptor –> target for beta arrestin (stops signaling )
beta arresting
stops signaling
Angiotensin receptor system
- binds to 7TM receptor
- activates G protein
- activates phospholipase C
- breaks down PIP2–> DAG and IP3
- DAG stimulates protein kinase A (w/ Calcium)
- IP3 stimulates release of calcium from ER–>calcium controls vessel contraction
- Calcium bind calmodulin
- CaM Kinases phosphorylate proteins.. .
Angiotensin
regulates blood pressure
PIP2
component of membrane
broken into DAG and IP3 by phospholipase C
DAG
2nd messenger
remains in or near lipid bilayer (nonpolar tails)
stimulates protein kinase A
protein kinase A stimulated by…
calcium and DHE together
located at cell membrane so can interact with DHE
IP3
2nd messenger
soluble in cytoplasm
binds to receptor in ER
opens channel and Ca from ER come out
Calcium…
“3rd messenger”
normally kept low to prevent it from binding proteins and precipitating DNA
essential for muscle contraction
EF Hands
structure in Calcium binding proteins
Calmodulin
calcium binding protein
keeps concentration low
able to bind other proteins –> CaM Kinases stimulated to phosphorylate proteins
Peptide Hormone
protein that acts as a hormone
ex: angiotensin, insulin, EGF
Insulin released in response to
increased glucose in blood, cells take it up
insulin receptor
not a 7TM, no G protein
in membrane of target cell
dimeric form
2 extracellular alpha subunits, 2 intracellular beta subunits
beta subunits of insulin receptor
have active site, tyrosine kinase
no insulin binding…
subunits are non-phosphorylated, kinase is inactive
binding of insulin…
moves units of dimer closer together –> phosphorylate tyrosine resides on each other –> activation
phosphorylated tyrosines on insulin receptor
bind IRS-1 via SH2 domain
tyrosines on IRS-1 phosphorylated as well
SH2 domain..
recognizes and binds phosphorylated tyrosines
phosphorylated tyrosines on IRS-1 bind
BOB
BOB…
PIP2–>PIP3 in membrane
PIP3 is binding target for
PDK1
PDK1
phosphorylates Akt
Akt
activates pathway –>GLUT4 to cell surface –>glucose into cell
EGF
stimulates cells to grow and divide
EGF receptor
monomer in cell membrane w/out EGF
binding of EGF to receptor…
receptor dimerize (interact with another receptor bound to EGF) phosphorylate each other at tyrosine
phosphorylated tyrosines of EGF receptor bind
Grb-2
Grb-2b binds
SOS
SOS…
interacts with Ras –> cell division stimulated
Ras and G proteins…
bad enzymes, hold onto GDP for few minutes before hydrolyze to GTP
on long enough to pass signal
oncogene
gene in which mutations can happen that lead to uncontrollable growth
proto-oncogene
normal, unmutated form of protein
important role in controlling cells
Ras
proto-oncogene
mutations interfere with ability to cleave GTP to GDP–> uncontrolled cell growth
Ras becomes as oncogene
src
proto-oncogene that participates in controlling cell’s decision to divide
first oncogene discovered: chicken with virus had cancer
when src tail are phosphorylated on tyrosine…
does not divide
when src tail are not phosphorylated on tyrosine…
stimulates cell division
viral form of src..
mutated such that does not have carboxyl tail…favors cancer bc cant be turned off (no tail to phosphorylate)
ABL
tyrosine kinase that plays role in controlling when cells divide
BCR-ABL
fusion made in greater quantities than ABL alone –>cells divide uncontrollably
BCR-ABL tumors treated with
Gleevec tyrosine kinase inhibitor
Her
related to EGF receptor
proto-oncogene
normally present in low amounts
bind to EGF receptor–>process
mutation in Her ..
favors Ras and cell division
Her and breast cancer
block with antibody –> stop process
Herceptin
negative delta G
goes forward
positive delta G
reverse favored
delta G = zero
in equilibrium
delta G zero
delta G measured under standard conditions, all products and reactants at 1M
delta G zero prime
prime to encompass aq. solutions at pH 7 for biological systems
as reactants increase…
products/reactants decreases
ln is negative
delta G is more negative
as products increase..
products/reactants increase
ln is positive
delta G is more positive
products/ reactants >1
ln is positive
products/reactants <1
ln is negative
electron carriers
NAD+/NADH
FAD/FADH2
NADP+/NADPH
6 classes of reactions catalyzed by enzymes
redox ligation isomerization group transfer hydrolytic lyases
primary way cells get energy
oxidation
glycolysis step 1
hexokinase
glucose + ATP –> G6P
negative delta G zero prime
glycolysis step 2
phoshpoglucoisomerase
G6P –> F6P
glycolysis step 3
PFK
F6P –> F-1,6 BP
negative delta G zero prime
most important step for regulation
glycolysis step 4
aldolase
F-1,6 BP –> DHAP and G3P
positive delta G zero prime, energetically unfavorable
1 6C molecules –> 2 3C molecules
glycolysis step 5
TPI (perfect enzyme)
DHAP –> G3P
at end of energy investment phase…
used 2 ATPs (steps 1 and 3)
make 2 G3Ps (steps 4 and 5)
glycolysis step 6
G3PDH G3P --> 1,3-BPG NAD+ --> NADH only redox reaction energy from oxidation of aldehyde --> acid
glycolysis step 7
phosphoglycerate kinase 1,3-BPG --> 3-phosphoglycerate (3PG) ADP --> ATP phosphate from 1,3-BPG to ADP to make ATP substrate level phosphorylation
