Biochem: Proteins 0814 And 15

Front 1

Macromolecules
Examples of small molecules

Back 1

Proteins, Nucleic Acids, Sugars, Lipids

Metals and Ions (Ca, Fe, Zn, Cl-...), Vitamins, Hormones
Nucleotide Triphosphates, Sterols

Front 2

4 levels of aminoacid interaction that make protein structure

Back 2

Primary structure
Secondary structure
Tertiary structure
Quaternary structure

Front 3

What is the predominant macromolecule in cells?

Back 3

Proteins are the predominant macromolecule in cells, comprising approximately 50% of their dry weight.

Front 4

What dictates the 3D structure of a protein?
What does the 3D structure dictate?

Back 4

Amino acid sequence and the characteristics of their side chains.
The ability to carry out a specific function.

Front 5

Give some examples of protein functions

Back 5

1. enzymes (catalyze biochemical reactions)

2. carriers of small molecules (such as oxygen, electrons, iron)
3. structural supports for cells and tissues, e.g. collagen, hair, feathers
4. movement (actin, myosin, etc.)
5. hormones (insulin, endorphins, nerve growth factor)
6. immune protection (antibodies/immunoglobulins)
7. specific responses to the environment (vision: rhodopsin absorbs light)
8. exotic roles (antifreeze in the blood of certain fish; tetanus toxin in bacteria)

Front 6

What are the 4 constituents covalently bonded to the alpha carbon in an amino acid?

Back 6

1. an amino group;
2. a carboxyl group;
3. a hydrogen atom; and
4. an R group (side chain)

Front 7

What version of amino acids are found in proteins?

Back 7

L-amino acids

Front 8

Describe a peptide bond

Back 8

Covalent bond formed between AAs, from the carboxyl group of one AA to the amino group of another. A water molecule is lost when this bond is formed.

Front 9

Which end is the starting end of an AA by convention? In which direction does ribosomal synthesis occur?

Back 9

The amino (N-terminus).

From N-terminus toward the C-terminus.

Front 10

Why is the peptide bond planar?

Back 10

The peptide bond is planer due to its partially double bond character and thus imparts some rigidity to the polypeptide chain.

Front 11

At Ph 7, which AAs are charged and polar?

Back 11

Glutamic acid, Apartic acid (carboxyl group is neg charged)
Lysine, Arginine, and Histadine (positively charged, considered basic AAs)
** energetically favorable for these AAs to be exposed to polar solvent

Front 12

At Ph 7, which AAs are uncharged and polar?

Back 12

Glutamine, asparagine
Serine, threonine, tyrosine (have hydroxl groups on side chain, so polar in part)
Cysteine (contains sulfur)
** energetically favorable for these AAs to be exposed to polar solvent

Front 13

At Ph 7, which AAs are nonpolar and some which are hydrophobic?

Back 13

Valine, leucine, isoleucine,
Phenylalanine and tryptophan (aromatic)
Proline (has special side chain bonding, so no amide proton)
**Glycine and alanine are neutral
**energetically UNfavorable for these AAs to be exposed to polar solvent

Front 14

Which AAs are aromatic?
Which has sulfur?
Which have hydroxyl groups?

Back 14

Phenyalanine, tyrosine and tryptophan
Cysteine, methionine
Serine, threonine, tryosine

Front 15

What are some post-translational modifications that can occur to AA side chains once in a polypeptide.
What kind of modifications are these?

Back 15

Phosphorylation, acetylation, methylation, glycosylation, prenylation, ubiquitylation, disulfide bond
Covalent modifications

Front 16

What is pKa?

Back 16

The pH at which a weak acid or base is 50% protonated.

Front 17

At high pH are side chains protonated or unprotonated?
At low pH?

Back 17

Unprotonated mostly.
Protonated mostly.

Front 18

What is driving force behind protein folding?

Back 18

Folded form is energetically more favorable.

Front 19

what does ΔG describe?

When a process is spontaneous the ΔG is what?

Back 19

free energy, whether a process will occur spontaneously.
Negative.

Front 20

What does positive enthalpy describe? negative?

What is entropy?

Back 20

If ΔH is positive, heat is gained/absorbed. If negative, heat is lost.
Entropy is the disorder of a system, an increase in disorder is energetically favorable.

Front 21

Secondary structure of proteins is what? What are some examples or regular secondary structures?

Back 21

local spatial arrangement of AA residues in relation to one another. Beta sheet and alpha helix.

Front 22

The 4 types of non-covalent bonds/interactions.

Back 22

Hydrogen bonds
(two electroneg atoms sharing a hydrogen atom)
Salt bridges (oppositely charged groups)
Van der Waals (attractions between adjacent non-repelling atoms)
Hydrophobic interactions

Front 23

Properties of Hydrogen Bonds in protein structure.

Back 23

Distance between donor and acceptor is fixed, according to the participants.
All three atoms must be linear due to partial covalent bond character.
Water can form h bonds with protein and mitigate the strength of the internal H bonds.

Front 24

Proteome Definition

Back 24

The term proteome is derived from
"proteins expressed by the genome." The genome (DNA) is the blueprint for an organism. By design, the genome is static and the proteome dynamic, constantly
being reshaped to meet the changing needs of a cell and/or organism.

Front 25

What techniques use solubility to isolate proteins?

Back 25

Salting out: The solubility of proteins varies with the ionic strength of the medium. Differential
precipitation can be accomplished by slowly increasing the salt concentration of the medium. isoelectric precipitation, Proteins will often precipitate out of solution at their isoelectric point
(pI)

Front 26

What is the isoelectric point(pI) of a protein?

Back 26

The pH at which there is no net charge on the protein.

Front 27

What technique separates proteins based on charge?

Back 27

Ion exchange chromatography - columns with a fixed negative (or positive) charge will bind proteins with a
patch of positive charge (or negative) on their surface. Bound proteins can be selectively eluted by a
gradual increase in the salt concentration, which breaks the salt bridges that form between the proteins
and the charge on the column. Since the pH of a solution affects the net charge on a protein, the pH of the solvent used to run an ion exchange column is a very important parameter.

Front 28

What technique uses binding affinity to separate proteins?

Back 28

Biospecific Chromatography-This technique depends on the biological activity of the protein.
Proteins can be purified to a large degree by chromatography on a column derivatized with a molecule
to which they bind with high specificity, such as a substrate-analog for an enzyme, a toxin, or a specific
inhibitor. Antibodies against a sequence from the protein of interest can also be made into a highly specific affinity resin.

Front 29

What is one of the techniques use size to separate proteins from small molecules?

Back 29

Dialysis – a way to separate proteins from small molecules, e.g., salt. Proteins (in a high salt buffer) are placed in a bag that has a semi-permeable membrane. The large protein molecules can not traverse the membrane, while the small molecules (salt) can. Dialysis is one way to lower the salt concentration of a protein
preparation. Lowering the salt concentration is an important prelude to ion exchange
chromatography.

Front 30

What is a dalton?

Back 30

One Dalton is equivalent
to the molecular weight of a hydrogen atom. Proteins range in size from molecular weight of a few thousand Daltons to complexes of several million Daltons.

Front 31

What is one of the techniques use size to separate large proteins from small proteins?

Back 31

Gel filtration chromatography - columns of material (beads with pores) that retard the mobility of small proteins
but not large ones. A sample, of a mixture of proteins, is
applied to the top of a column. Small molecules can enter the beads, while
larger molecules can not. As a result, the smaller molecules take longer to flow through the column; and the larger molecules come through the column sooner.

Front 32

What is one of the techniques use size to separate proteins based on mass?

Back 32

Proteins are denatured and the anionic the detergent used binds to the amino acids, giving the protein a net negative charge roughly proportional to its mass. The proteins are then separated by their size by electrophoresis. The small proteins move through the gel more rapidly than the larger ones. Proteins are visualized by staining.

Front 33

What are peripheral membrane proteins?

Back 33

Peripheral membrane proteins are associated with the surface of the membrane and associate
with both the polar head groups of the lipids and the integral membrane proteins through ionic
interactions. These proteins can be removed from the membranes by mild extraction with high salt or
pH. Once extracted, they behave as typical soluble proteins in aqueous solution.

Front 34

What are intergral membrane proteins? How are they extracted from the protein for analysis?

Back 34

Integral membrane proteins have a segment of their sequence directly inserted into the hydrophobic region of the lipid bilayer in contact with the fatty acid chains of the membrane lipids. Integral membrane proteins are only extracted from the membrane using a detergent to disrupt the membrane. Detergents displace the membrane lipids from the hydrophobic surface of the protein. Once
removed from the membrane, integral membrane proteins will aggregate due to their hydrophobic regions unless lipids or detergents are present.

Front 35

What are two methods of protein identification?

Back 35

The traditional way (protein sequencing) and the new methodology (mass matching) are used when one isolates a protein according to its
function (i.e. binds to a drug) and one wants to know the polypeptide sequence of that protein.

Front 36

How does protein sequencing work?

Back 36

EDMAN DEGRADATION an automated process that sequentially removes one labeled amino acid at the N-terminus of the peptide without disrupting the other peptide bonds.

Front 37

How does mass matching work?

Back 37

The isolated protein is digested with trypsin and the mass of the peptides generated will be measured by mass spectrometry.
Second, using the genome of the organism you are studying a hypothetical list of all proteins and their amino acid sequence is constructed and a computer theoretically digests them and generates the possible peptides and masses. The actual and theoretical peptides are then matched.
Using more sophisticated versions of this technology, methods have been devised to
identify hundreds of proteins and their relative quantities in a complex mixture (i.e. an organelle or pathogenic organism).

Front 38

What are some of the uses for the info in protein and gene databases of amino acid sequences?

Back 38

Can see if similarities exist, if protein belongs to an established family.
See signaling sequences for destinations or control mechanisms.
Provides info for preparing antibodies against protein of interest.