BB450 exam 1 Flashcards
DNA bases and how they bind
adenine - thymine (2 H bonds)
guanine - cytosine (3 H bonds)
acid
anything that can give a proton
salt
something that has lost a proton
Ka =
[H+][A-]/[HA]
pKa
-logKa
constant for an acid, strength
lower pKa
stronger acid
higher pKa
weaker acid
extend of dissociation of weak acid related to…
pH of solution
by Henderson - Hasselbalch equation
Henderson - Hasselbalch equation
pH = pKa = log [A-]/[HA]
ratio of salt to acid increases…
pH increases
ratio of salt to acid decreases
pH decreases
equal amounts of salt and acid…
log of 0 = 1
pKa = pH where salt = acid
Le Chatelier’s principle
system responds to disruption by trying to reestablish equilibrium
if add HCl…
A- will bind to them
decrease A- and increase HA, driven to left
if add NaOH…
OH- will bind to H+ to form water
protons needed from HA, driven to right
HA decreases, A- increases
buffer will be at maximum capacity when…
[HA] = [A-] acid = salt
buffering range
+ or - 1 pH unit from pKa
buffer maximally effective when..
pH = pKa
if add HCl in buffer region…
each HCl molecule –> one mol/molecule salt to convert to 1 mol/molecule of weak acid
if add NaOH in buffer region…
each NaOH molecule –> one mol/molecule acid to convert to one mol/molecule of salt
estimating charge…
pH is 1 or more units above pKa - proton OFF
pH is 1 or more units below pKa - proton ON
pI
pH at which charge is 0
average of 2 pKa values where charge of aa is 0
pKa describes pH at which…
1/2 of each protons are off and 1/2 are on
almost all amino acids exist is ___ form
L (rarely D)
5 groups of amino acids
aliphatics hydrophobics polar positive R group ( R amine) negative R group (R carboxyl)
Aliphatics
glycine alanine proline valine leucine
hydrophobics
isoleucine
methionine
tryptophan
phenylalanine
polar
serine threonine tyrosine asparagine glutamine cysteine
positive R-group
lysine
arginine
histidine
negative R- group
aspartate
glutamate
glycine
smallest R group - H
most flexible
proline
R group connects with amine –> ring
can’t rotate, very inflexible
–> bends in protein structure
only favors trans peptide bond 100:1 (not 10,000:1)
methionine
C-S-C (not very reactive)
first aa in virtually every protein
cysteine
sulfhydryl S-H (very reactive)
reacts with other cysteines –> disulfide bond
peptide bond
between carboxyl on left, amino on right
phi
rotational angle between alpha amino and alpha carbon
psi
rotational angle around alpha carboxyl and alpha carbon
secondary structure
regular repeating structures arising from interactions between amino acids less than 10 apart
proteins with only primary and secondary structure
fibrous proteins
ex: collagen
separating large from small molecules
dialysis
gel filtration/gel exclusion
separate charged molecules
ion exchange chromatography
cation exchange or anion exchange
separate proteins that bind to a specific molecule
affinity chromatography
separate molecules on basis of polarity
HPLC (high performance liquid chromatography)
separate negatively charged molecules by size
agarose gel electrophoresis
SDS - PAGE (polyacrylamide gel electrophoresis)
separate by pI
isoelectric focusing
2D gel electrophoresis
isoelectric focusing (by pI) then SDS - PAGE (by size)
MALDI - TOF
determine molecular weight of large molecules
by time it takes volatized crystal to hit electrode
cleaving agents
cyanogen bromide
trypsin
chymotrypsin
carboxypeptidase
cyanogen bromide
cleaves carboxyl side of methionine
trypsin
cleaves carboxyl side of lysine and arginine
chymotrypsin
cleaves carboxyl side tyrosine, tryptophan, phenylalanine, leucine, methionine
carboxypeptidase
cleaves amino side of carboxy-terminal aa
DDT
reduces SS to SH
heme
protoporphyrin IX complexed with iron
iron held in place by
4 N from protoporphyrin IX and a histadine
form of iron involved in carrying oxygen
Fe2+ ferrous
T state
releases O2
low O2 affinity
R state
bind O2
high O2 affinity
bind of O2 help flip from ___ to ___
T to R
release of O2 help flip from __ to __
R to T
binding of 2,3-BPG converts hemoglobin from ___ to ___
R to T (releases O2)
fetal hemoglobin exists more in
R state (doesn’t release O2 as readily)
____ favors release of oxygen
CO2, 2,3- BPG, protons
rapidly metabolizing tissues generate…
protons, lower pH
negative delta G
reaction goes forward
positive delta G
reaction goes backwards
delta G = 0
at equilibrium
plot of velocity vs. substrate concentration
hyperbolic plot
Vmax
when enzyme is saturated with substrate
depends on amount of enzyme
Kcat
# molecules of product made/molecule of enzyme/1 sec turnover number constant for enzyme
Km
substrate concentration that give Vmax/2
affinity of enzyme for substrate
constant for enzyme
higher Km…
lower affinity
need more substrate to reach Vmax/2
lower Km…
higher affinity
need less substrate to reach Vmax/2
equilibrium
forward reaction = reverse reaction
lineweaver- Burk plots
double reciprocal plot
1/V vs. 1/[S]
y intercept of lineweaver - burk
I/vmax
x intercept of lineweaver - burk
-1/Km
slope of lineweaver - burk
Km//vmax
competitive inhibition.. Vmax
does not vary
competitive inhibition… Km
does vary (need more substrate to get to same velocity)
non-competitive inhibition … Vmax
lowered
non- competitive inhibition… Km
unchanged (increased substrate cannot eliminate effect of inhibitor)
Forces that stabilize tertiary structure
H bonds Disulfide (cysteine) Ionic Hydrophobic interactions Metallic
Polar and charged on ___
Outside
Interacting with water to dissolve
Non polar and uncharged on___
Inside
To avoid water
Pourin
In cell membranes
Allow water to pass through
Polar, charged on inside
Beta-mercaptoethanol
Disulfide to sulfhydryl
SS- SH SH
Urea
Destroys H bonds
Affects 2, 3, and 4 structure
DTT
Reduces disulfide to sulfhydryl bonds
Guanidium chloride
Denatures H bonds
