enzymes Flashcards
oxidoreductases
REDOX rxns that involve the transfer of electrons
transferases move a
function group from one molecule to another
hydrolases catalyze
cleavage w/ the addition of water
lyases catalyze cleavage without
the addition of water and transfer of electrons
isomerases catalyze the interconversion of
isomers
including constitutional isomers + stereoisomers
ligases join two large
biomolecules often of the same type
lipases catalyze the
hydrolysis of fats
dietary fats are broken down into
fatty acids and glycerol/other alcohols
kinases add a
phosphate group
type of transferase
phosphatases remove a
phosphate group
type of transferase
phosphorylases introduce a
phosphate group into an organic molecule (notably glucose)
exergonic rxn
RELEASE energy
gibbs free E = negative
endergonic rxns
require energy
gibbs free E = positive
enzymes do not alter the
gibbs free E or enthalpy or the final equilibrium position
enzymes only alter the
changing the rate of the reaction
as substrate increases, so does the
rxn rate until a max value is reached
K is the [S] at which an enzyme…
runs at half its Vmax
Vmax is the max rate at which an enzymes can….
catalyze a rxn
all enzyme active sites are saturated w/ substrate
cooperative enzymes display what kind of curve
sigmoidal curve b/c of activity change with substrate binding
enzymes stabilize the
transition state
enzymes provide a favorable microenvironment and/or…
bonding w/ the substrate molecules
site of catalysis is called
the active site
lock and key theory
enzyme and substrate are complementary and fit together like a key into a lock
induced fit theory is where the enzyme and substrate…
undergo conformational changes to interact fully
metal cation that is required by some enzymes
cofactor
what molecule is required by some enzymes
organic molecules
how does temp and pH affect enzymes
denaturation and loss of activity due to loss of 2/3/4 structure of proteins
salinity can impact the actions of
enzymes
feedback inhibition is when an enzyme is inhibited by
high levels of a product from later in the same pathway
reversible inhibition is the ability to replace the inhibitor with a….
compound of greater affinity or to remove i.t. using mild lab treatment
competitve inhibition
when inhibitior is similar to the substrate
binds at the active site
blocks substrate from binding
competitive inhibition can be overcome by
adding more substrate
vmax and km in competitive inhibtion
vmax unchanged
km increases
uncompetitive inhibition is when the inhibitors binds…
only w/ the ES complex
vmax and km in uncompetitive inhibtion
vmax and km decrease
noncompetitive inhibition is when the inhibitors binds with…
equal affinity to the enzyme and ES complex
vmax and km for noncompetitive inhibition
vmax decreases and km unchanged
mixed inhibition is when the inhibitor binds with unequal…
affinity to the enzyme and ES complex
vmax and km in mixed inhibition
vmax decreases
km increases/decreases depending on if the inhibitor has a higher affinity for the enzyme or ES complex
irreversible inhibition alters the enzyme in such a way that the
active site is unavailable for a prolonged duration / permanently
suicide inhibitor is a substrate analogue that binds…
IRREVERSIBLY to the active site via a covalent bond
allosteric effector binds at the allosteric site and induces a…
change in the conformation of the enzyme
the substrate can no longer bind to the active site
allosteric effector displays
cooperativity
does not obey MM-kinetics
positive allosteric effectors exert a
positive effect
increase activity
negative allosteric effector exert a
negative effect
decrease activity
homotropic effector is an allosteric regulator that is also the…
substrate
e.g. O2 is a homotropic allosteric regulator of hemoglobin
heterotropic effector is an allosteric regulator molecule that is
different from the substrate
what is phosphorylation
covalent modification with phosphate
catabolism
phosphorylated = active
anabolism
phosphorylated = inactive
glycolsylation is the covalent modification with
carbohydrates
zymogens is the
precursor to an enzyme
zymogens are secreted in an inactive form and are…
activated by cleavage
which proteins made up the cytoskeleton/anchoring proteins/EC matrix
structural proteins
common structural proteins
collagen
elastin
keratin
actin
tubulin
motor proteins have 1+ heads capable of
force generation through a conformation change
motor proteins have catalyic activity, acting as
ATPases to power movement
common applications w/ motor proteins
muscle contraction
vesicle mvmt within cells
cell motility
e.g. myosin, kinesin, dynein
binding proteins bind a
specific substrate
either to sequester it in body or hold concentration at steady state
what does cell adhesion molecules allows cells to do
allows cells to bind to other cells/surfaces
cadherins are
calcium-dependent glycoproteins holding similar cells together
intergrins have two membrane-spanning chains and allow cells to…
adhere to proteins in EC matrix
selectins allows cells to adhere to
carbs on surfaces of other cells
commonly used in immune system
antibodies are aka
immunoglobulins (Ig)
used by immune system to target a specific antigen
ion channels can be used for
regulating ion flow in/out of cell
ungated channels are
ALWAYS OPEN
voltage-gated channels are
OPEN within a range of membrane potentials
ligand-gated channels open in the
presence of specific binding substance (usually a hormone or neurotransmitter)
enzyme-linked receptors participate in cell signalling through
EC ligand binding and initiation of 2nd messenger cascades
GCPR has a membrane-bound protein called the
G-protein (has three subunits)
1st messenger ligand initiates the
2nd messenger and the cascade response
epinephrine is a ligand
2st messenger
at the end of the GCPR process,
phosphodiesterase deactivated cAMP
GTP hydrolyzed back to GDP
electrophoresis uses a
gel matrix to observe migration of proteins in response to an electric field
native PAGE maintains the protein’s shape but results are
difficult to compare b/c mass/charge ratio differs by protein
SDS-PAGE denatures proteins and masks the
native charge so size comparison is more accurate
func protein cannot be recaptured from gel
isoelectric focusing separates proteins by the
pI
protein migrates towards electrode until i.t. reaches region where pH = pI of protein
chromatography separates protein mixtures on the basis of
their affinity for a stationary phase or mobile phase
column chromatography uses beads of a
polar compound (stationary phase) with a non-polar solvent (mobile phase)
ion-exchange chromotography uses a
charged column and a variably salin eluent
size exclusion chromotography relies on
poroous beads
larger molecules elute first - not trapped in small pores
affinity chromotography uses a bound receptor or ligand and an eluding with a….
free ligand or receptor for the protein of interest
how is protein structure primarily determined
XRAY crystallography after protein isolated then NMR can be used
edmand degradation determines
AA sequence
bradford protein assay is most
common
uses color change from brown-green to blue
what does topoisomerase do
unwind the DNA double helix
helicase function
breaks H-bonds b/w the nitrogenous bases in order to separate DNA strands
single strand binding protein (SSB) binds to
single-stranded DNA and prevents annealing of ssDNA into double-stranded DNA
what does DNA primase do
catalyzes synthesis of RNA primer
RNA primers allows for
DNA replication to start
short RNA nucleotide sequences that are complementary to the ssDNA
DNA polymerase adds
nucleotides to the growing strand
how to read DNA template
3’ to 5’
how is the new strand of DNA synthesized
5’ to 3’
DNA polymerase removes the
RNA primer at the end of the strand
okazaki fragments are short, newly synthesized DNA fragments formed
on the lagging template strand during DNA replication
DNA ligase join the
DNA strands together by catalyzing formation of phosphodiester bonds
proofreading DNA/RNA
DNA has proofreading, RNA does NOT
DNA replication is more (what) than RNA replication
accurate
stability of DNA/RNA
RNA is less stable b/c it contains sugar ribose instead of DNA’s deoxyribose
because RNA is less stable, it…
degrades rapidly in cytoplasm
strands of DNA wrap around a
histone protein forming nucleosomes
nucleosomes coil together and form
chromatin
chromatin loops and coils together to form
supercoils
supercoils bunch together to form
chromosomes
charge of DNA
negative due to phosphate backbone
3 parts of a nucleotide
5 carbon sugar
nitrogen-rich base
phosphate group
nucleoside lacks
phosphate group
nucleotide pairs
adenine - thymine
guanine - cytosine
adenine-thymind has how many H-bonds
2 H-bonds
guanine-cytosine has how many H bonds
3 H bonds
stronger than A-T
what holds DNA backbone together
phosphodiester bonds between sugar and phosphate groups
hydrogen bonds hold what together in DNA
nucleotide bases together inside the double helix
how many rings in pyrimidine
1 ring
how many rings in purines
2 rings
pyrimidine ring fused to an imidazole ring
pyrimidines
cytosine
thymine
uracil (RNA only)
purines
adenine
guanine
purines and pyrimidines have a 1:1 ratio as per
Chargaff’s rule
purine and pyrimidine have what kind of width
UNIFORM
purine + purine is too
wide
pyrimidine + pyrimidine is too
narrow
diameter of DNA double helix =
20 angstroms
RNA is a polymer of
nucleotides
how does RNA differ from DNA
1) RNA is single stranded
2) sugar is ribose + more reactive
3) nitrogenous base is uracil (U) not thymine (T)
