Exam 1 Flashcards
(141 cards)
1
Q
Three domains of life and order of relationship
A
Bacteria, archaea, then eukarya
2
Q
aerobic vs anaerobic
A
aero= O2 and ana is no O2
3
Q
Chemotrophs
A
Oxidize chemical fuel
4
Q
Lithotrophs
A
oxidize inorganic fuel such as iron and sulfur
5
Q
organotrophs
A
oxidize organic fuels
6
Q
most of the dry mass of any cell is composed of
A
H, C, N, O
7
Q
sulfhydryl
A
R-S-H
8
Q
thioester
A
R-C(=O)-S-R
9
Q
C(=O)-O-
A
carboxylate
10
Q
R-O-R
A
ether
11
Q
R-C(=O)-O-R
A
ester
12
Q
RNA is built from monomers called:
A
nucleotides
13
Q
basic biochemical building blocks
A
amino acids, nucleotides (nuclei acids), lipids(lipid bilayer), sugars (cellulose)
14
Q
chiral molecule
A
rotated molecule cannot be superimposed on its mirror image
15
Q
achiral molecule
A
rotated molecule can be superimposed on its mirror image
16
Q
enantiomers
A
mirror image
17
Q
diastereomers
A
non-mirror images
18
Q
racemic mixture
A
mixture with equal concentrations of enantiomers
19
Q
R (CW) is:
A
D
20
Q
S (CCW) is:
A
L
21
Q
bond angle between tetrahedral carbons
A
109.5
22
Q
why dont double bonds rotate
A
the interconversion requires bond-breaking that enzymes can accomplish, but cannot happen spontaneously at low temp
23
Q
supramolecular to macromolecules to monomeric subunits
A
- chromatic-DNA-nucleotides
- plasma membrane- protein- amino acids
- cell wall-cellulose-sugars
24
Q
dynamic steady state
A
Inputs of energy and fresh material are regularly required to produce and maintain molecules of life and for reproduction of living organisms. Implies continual input of new material and energy as well as continual discharge of waste products and heat energy.
25
gibbs free energy equation
change G= change H-T (change S)
-H= releases heat
+S- increases randomness
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free energy
amount of energy available to do work, determines thermodynamic favorability
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exergonic
reaction in which the products have less free energy than reactants-releases net free energy (negative G)
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endergonic
reaction in which the products have more free energy than reactants-requires input of free energy (positive G)
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enthalpy
(H) heat content of reacting system-relfects the number and kinds of chemical bonds involved
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entropy
S- disorder of system (number of molecules, freedom to move)
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actual vs standard free energy change
actual is system function of actual conditions, while standard represents standard conditions (constant)- 298K, 1M reactants, 1atm P
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benefit of coupling reaction
two or more reactions may be coupled, allowing a new unfavorable reaction to be part of a favorable sequence of reactions and thus to be possible and efficient
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what do enzymes do?
they lower the activation energy
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transition state
state generated by breaking bonds
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difference between functional components in DNA and RNA
RNA has an extra -OH group so it is ribose instead of deoxyribose
36
what does together in DNA
AT then GC, B form helix
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central dogma
DNA transcribed to messanger RNA (job site blueprint) which is translated to polypeptide chain (unfolded hexokinase- inactive protein) the is folded into native structure of helokinase (catalytically active hexokinase- fully functional protein)
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what determines cellular function
it is conferred by structure, not sequence alone
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to get mutation in protein?
two rare mistakes- one is DNA replication and second rare mistake on second hexakinase gene
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Miller and Urey, 1953
Abiotic production of biomolecules, established conditions for abiotic production of biomolecules in fairly short time
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endosymbiont conjecture
evolution of eukaryotes into nonphotosyn and photosynthetic (cyanobacterial genome)
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hydrogen bond
An electrostatic interaction due to unequal charge distribution. Three molecules in linear form- most stable/strong. Atom with H attached is the donor.
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amphipathic
substances with both polar and non-polar regions
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water forms weak interactions with solutes including: (weak compared to covalent bonds)
hydrogen bonds, van der waals, hydrophobic effects, ionic interactions
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ionic interactions
attraction (bond) between oppositely charged ions or molecules. can be attractive or repulsive.
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hydrophobic effect
Entropy based favorable free energy effect for close packing of hydrophobic molecules
non polar substances do not dissolve in polar solvents to any significant extent. So water forms "cages" of highly ordered water around non-polar molecules (decreasing entropy of water phase). As hydrophobic molecules enter water, the water surrounding them must become ordered- unfavorable. but putting them in cages, less water them so more favorable. Driving force is increased water entropy.
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micelles
all hydrophobic groups are sequestered from water, ordered shell of H20 molecules is minimized, and entropy is further increased.
48
van der waals interactions
Random variations in electronic clouds of any two atoms can create a transient dipole, which leased to a weak attractive force that brings the two atoms closer together. At the point where the attractive force is maximal (van der waals radius). Resonsible for atomic close packing. Individaully weak but very numerous.
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condensation reaction
eliminates water and is usually endergonic
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hydrolysis reaction
adds water to break a bond and is usually exergonic
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ionization
gaining or losing an electron to form ions
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reaction equilibrium (equilibrium constant= Keq)
when the rates of product and reactant formations become equal
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Kw (ion product of water)
1.0*10^-14 M^2
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pH of physiological pH
7.4
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Ka (acid dissociation constant)
Is a constant, a fundamental property of the acid. It doesnt change with changes in pH. It depends on the chemical structure and environment of the acid group.
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equivalence
complete deprotonation has occurred, meaning amount of added base is equal to the total number of ionizable protons in the weak acid. This can be used to determine the concentration of the weak acid.
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inflection point
observed when pH=pKa for the acid. At the inflectin point [HA]=[A-] and pH changes most slowly with added base near this pH. the midpoint of titration.
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effective buffering range
+/- 1 from inflection point (pKa)
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buffers
substances that can be added to aqueous systems to help resist overt changes in pH as acids or bases are added. Extremely important to control pH of cellular systems, as many near specific pH values.
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maximum buffering capacity
occurs at the pKa of weak acid/conjugate base system
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non-essential AA
```
Can be synthesized by mammals. Includes:
alanine
asparagine
aspartate
glutamate
serine
```
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Conditionally essential AA
Needed for rapid growth. Can be made from other 2 groups.
| Includes: Arginine, cysteine, glutamine, glycine, proline, tyrosine
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essential AA
Only obtained through animal diet. Must be supplied daily but can be found in muscle if necessary. Includes:
```
Histidine
Isoleucine
leucine
lysine
methionine
phenylalanine
threonine
tryptophan
valine
```
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zwitterions
dipolar ions and can act as acids (proton donors) or as bases (proton acceptors)
amino acid good example (when carboxylic is neg and amino is positive)
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difference between l and D
They have identical physical and chemical properties except for rotation of polarized light. They have different biological properties.
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Methionine
Initiator AA for protein synthesis. Also incorporated within proteins.
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maple syrup urine disease
Branched chain ketoaciduria. Defect in an enzyme that breaks down branched chain AAs. Results in toxic buildup of these AAs. Most notably Leucine and patients produce urine that smells like maple syrup. Rare trait.
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precursor to Tyr
Phenylalanine
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precursor to chatecholamines (dopamine, Nre, Ep)
Tyr
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AA that can H-bond
Tyrosine (Y) and tryptophan (W)
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precursor to serotonin
Trp
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PKU
Defect in the metabolism of phenylalanine. Leads to the buildup of metabolites. Extremely painful condidtion. Severe sequalae are like MSUD. Can be controlled by dietary restrictions (avoid artificial peptide sweeteners bsc they are converted into Phe after consumption)
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absorbances of tryptophan and tyrosine (aromatic side chains absorb light)
tyrosine: 274
tryptophan: 280 (extreme)
phenylanaline 257
larger side chain, large UV absorbance per mole
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forms disulfides which are nonpolar, and can stablize some structures
cysteine
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when does S- ionize?
8.2
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disulfide bond (cross link)
two adjacent sulfydryls can be oxidized to form a covalent bond. Is nonpolar, hydrophobic, and often structure stabilizing.
77
how many h bonds can asparagine and glutamine make
2
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pI (isoelectric point)
pH at which net charge is zero
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peptides
simply chains of amino acid residues linked by amide bonds made by condensation reactions between carboxylic acid and amino groups of amino acids
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peptide bonds
amide bonds made by condensation reactions between carboxylic acid and amino groups of amino acids. they are polar but not charged. The charged groups have vanished.
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what breaks disulfide bonds
DTT or B(2)-mercaptoethanol
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edman degradation
Usual strategy is to cleave with a few different reagents, sequence several peptides each by edman degradation and identify overlapping sequences to determine the entire protein sequence
83
collagen
connective tissue. 3 left handed helical chains are intertwined with interchain h bonding. Repeating sequence: G-X-Y. X is often proline and y is 4 hydroxyproline (modifies P to hydroxyproline). Production of hydroxyproline required for normal stability of collagen, failure of modification leads to serious connective tissue, abnormality; vitamin c required which leaves to the sever lack of vitC (scurvy)
84
D- AA
used in medicine
long lived in body (immune to degragation by enzymes)
contains same functional groups and same chemistry as normal r groups
great mimics
made synthetically
not cleaved by proteases
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peptide group
contains 6 atoms
| -C-CO-NH-C-
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omega (w)
close to 180 when trans or 0 when cis. between carbon and N (no rotation )
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phi bond (circle with line through it)
N and alpha carbon
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psi W with line
carbonyl and carbon
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ramachandran plots
plot of observed phi and psi angles showing sterically allowable configurations. (corresponds to secondary structures)
135-(-135)= b strands
-90 and 135 (right handed alpha helix
45 to 45 left handed alpha heliz
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secondary strucutre
local spatial arrangements of groups of amino acids
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good AA for alpha helices
Ala
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bad AA for alpha helices
pro and gly (high free energy cost)
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amphipathic alpha helices
hydrophilic on one face and hydrophobic on the other
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type I turn
has proline at 2nd position. (rigid backbone held in cis configuration conducive to reversing chair direction)
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type II turn
Has glycine at 3rd position (smallest side chain, thus backbone confirmation very flexible.)
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random coil
nonrepetitive but ordered secondary structural elements,
"not a regular repititive secondary structure"
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loops
are needed to connect secondary structure elements. typically 2-16 residues in length, and are important to function
98
most important bonds in native conformations>?
non covalent interactions (only marginally stable but still net favorable free energy of folding is small)
99
tertiary structure of protein
include the long range details of the AA positions, 3D structure (made from a variety of motifs and domains)
overall folding of a single polypeptide chain into its native 3D structure
100
motifs (supersecondary structure)
- distinct folding patterns that consists of one or more secondary structural elements
- stabilized by H-bonds, ionic interactions, van der waals, and hydrophobic effect
- can be amphiphatics
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domains
- independently folded and sometimes independently functional parts of a protein (fold up by itself)
- retain structure and function sometimes even in the absence of the rest of the protein
- separate domains can provide specific functions, can associate with each other or ligand and may be connected by linker
102
quaternary structure
consists of two or more separate polypeptide chains associating with each other in a specific manner that is required for optimal biological activity. (can be called dimer or trimer)
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Levinthal's paradox
protein folding cannot possibly operate as a random trial and error process. Instructions must be build into the proteins; such instructions represent information coded in sequence that specifies a folding pathway.
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refolding in anfinsen's experiment
AA sequence must have controlled correct folding of tertiary enzyme structure
105
amyloid fibrils
Misfolded structures alter their proper configuration such that they erroneously interact with one another forming insoluble fibrils. -> leads to cell death
106
chaperones
assist many proteins to fold correctly in the cell
107
conjugated proteins
many proteins have some non-amino acid components
ex. serine can be phosphorylated into phospho-serine and now have a large neg. charge
108
AA that can be modified
serine, lysine, threonine, asparagine
109
modifications at n-terminus
Myristoylation (adding a FA to protein affects its subcellular localization). Important in targeting proteins to membranes. Irreversible addition- FA's very hydrophobic
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modifications to C-terminus and cysteines
Palmitoylation (cellular machinery discriminates based on size of FA added to protein or site of FA modification). Targets proteins to membranes. Reversible modification. Aids in trafficking proteins to appropriate vesicles and organelles.
111
signal peptide
stretch of hydrophobic residues near N-terminus may target protein for export from cell or to intracellular membrane location. Has a cleavage site that is cleaved by signal peptidase so it adopts correct, new fold.
112
Nuclear localization signal (NLS)
AA sequence that targets transport of eukaryotic proteins from the cytosol into nucleus (-PKKKRKV-)
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Binding motif
bind and cleave target
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what do you know if proteins have similar protein sequences
they are likely to have similar 3D structures ( and maybe similar functions)
115
what mutation occured in sickle cell patients
E6V (Glu to Val at 6th position)
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Exploit the differences in proteins based on:
- solubility in salts or solvents
- net molecular charge
- surface regions of +/- charge
- overall shape and size
- hydrophobic interactions
- specific interactions with ligands
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first three steps in protein purification
1. choose source material
2. choose a buffer
3. Homogenization or differential configuration (sep. soluble and nonsoluble parts)
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four types (of fractionation of extract) methods
1. precipitation
2. chromatographic
3. centrifugation
4. electrophoresis`
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Ion exchange chromatography
- used early in whole purification
- scalable, ion exchange resins cheap, high capacity, and can be fast
- use either an anion or cation exchange with NaCl (charged at all pH) that are attached to beads
- protein replaces Na+/Cl-, but the reversed with high conc. of NaCl.
- depends on the net charge of the protein molecule (dont want it at pI of desired protein)
120
pH < pI
protein is neg charged
121
pH> pI
protein is positively charged
122
gel filtration (gel permeation) chromatography
Protein molecules separate by size. Larger molecules pass more freely, appearing in the early fractions. Input must be small soluble conc. protein fraction as eluted volume. Large proteins elute early (go around beads) and smaller proteins elute later (see entire tube as volume which they travel through beads). Based on size AND shape (rod vs sphere). Rod will act like a large sphere and wont enter holes). Beads are a neutral sugar polymer with lots of space in them. Good at end of sep. and volume must be 1/10 of total volume of column.
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Affinity chromatography
Protein mixture is added to column containing a polymer bound ligand specific for protein of interest. Input must be soluble protein fraction almost free of interfering activities often used as late step in process. Uses principle of affinity. Unwanted proteins are washed through column and protein of interest is eluted by ligand solution. Affinity method most selective of common purification strategies. Specific interactions with functional groups immobilized on beads. Tags can also be used in this process. ex- his-6.
124
SDS polyacrylamide Gel Electrophoresis
Migration rate depends on MW not on AA composition of folded shape. Can compare Mr standard to unknown protein. Each polypeptide runs as a discrete band, detected by non-specific stain to see nearly any protein. Complex proteins fall apart into their separate polypeptides. Can make a graph comparing MM (kDa) vs relative mobility to determine MW of unknown.
125
SDS
binds avidly to most proteins at fairly consistent mass ratio. 1.4mg SDS to 1 mg protein. 1 SDS per 2 AA residues. Causes proteins to unfold into random coil. All have high neg charge and constant diameter and similar shape. Only different is MW of protein. SDS is an artificial fatty acid analog. Binds to hydrophobic part and unfolds protein.
126
Native gel electrophoresis
- estimate native Mr but remember it may not be accurate since charge, size, and shape influence migration (as for gel filtration)
- useful mostly for looking at protein: protein interactions
- protein complexes
- protein/DNA or RNA and look how protein interacts with them (migrate differently so can observe shifts from known DNA)
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sedimentation analysis
- large particles sediment more rapidly than small particles, assuming same shape and same frictional coefficient
- spherical particles sediment more rapidly than another shape of same mass and density
- often encounter "s values" in older literature
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Bouyant density
- useful for organelles and particles
- sediment to an equilibrium bouyant density in density gradients
- can be made from salts (CsCl- very dense) or another material
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isoelectric focusing and 2-D gel electrophoresis
Isoelectric (1-D)- gel solution has a linear pH gradient (ampholytes). Proteins migrate in electric field based on charge. Each focuses at own pI. Final position based on pI and native complex can remain intact.
2-D: Turn gel tube 90 degress and separate in denature and reducing condition(SDS-PAGE and B-ME). Migration based on polypeptide size (Mr).
130
RBC
- 6 um
- filled with hemoglobin and metabolic enxymes
- lack nuclei and membranous organelles (increased SA with concavity)
- 150g/L of hemoglobin in blood
- HbA bears conc of 10mM
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myoglobin
cells that use large amounts of oxygen contain this protein (particularly in muscles). REPRESENTS O2 in STORAGE.
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Heme
- has four pyrrole rings linked by methene bridges (porphyrin) plus one Fe2+ metal ion---prosthetic group
- free heme binds strongly to molecular oxygen (oxidized to Fe3+)
- extends ligand binding ability of protein and provides a special functionality
133
prosthetic group
a compound that is permanently with a protein and contributes to the usual function of that protein (ex heme)
134
isosbestic points
-where spectra overlap (800 and 600). Can estimated amount of Hb oxygenated (HbA)
135
color of oxygenated vs deoxygenated hemoglob
de- dark purple (absorbs red)
| oxygenated- red (red isnt absorbed)
136
type of curve for myoglobin
hyperbolic, each O2 binds independently
137
higher Kd?
lower affinity
138
higher Ka?
higher affinity
139
T- state
- heme Fe doesnt fit well into porphyrin ring, so porphyrin flexed
140
R- state
-heme Fe is drawn into the plane of porphyrin, so ring is relaxed (causes movement of helix triggered by shift of heme Fe which triggers conformational change in Hb protein tetramer)
-O2 has higher affinity for R (versus T)
-
141
Selective pressure
Allows organisms to evolve new gene to adapt to changes in their environment
