Background Knowledge and Methodology Terms (BIO320)

A molecule that is capable of releasing or donating a hydrogen ion is termed an:

acid

A molecule that is composed of a central carbon surrounded by an amino group, a carboxyl group, a hydrogen atom and a variable side chain is called _____________________.

amino acid

antibody

A protein that binds to antigens

antigen

A molecule that elicits an immune response. Usually about 5-12 amino acids long.

_______ has an amide R-group that can be modified by glycosylation.

Asn

These amino acids have an amide R group and are NOT charged.

Asn & Gln

These amino acids have a carboxyl (-COOH) R group.

Asp & Glu

A molecule that is capable of accepting a hydrogen ion is termed a ____

base

beta-mercaptoethanol

A reducing agent that breaks the disulfide bonds between two cysteine residues.

Chase

The period of time during which non-radioactive amino acids are provided to a cell

Coimmunoprecipitation

Method in which an antibody is used to purify one protein and any other protein(s) associated with it.

Coomassie blue

General protein stain used to visualize proteins separated on a gel.

Two atoms share electron pairs when they have a __________ bond.

covalent

Strongest chemical bond

Covalent bond

This amino acid can form disulfide bonds

Cysteine

Negatively charged amino acids in single code

D & E

Detergent

A molecule able to disrupt all but covalent interactions

Differential centrifugation

Method used to separate different components of a cell by size.

Formed between two cysteine amino acids.

Disulfide bond

This bond can be broken by beta-mercaptoethanol or DTT.

Disulfide bond

Domain

Portion of a protein that folds independently of the rest of the protein.

Epitope

Small region of a protein where an antibody binds

what does a hydropathy plot do?

gives you hints about possible transmembrane domains

Immunoprecipitation

Method used to isolate a specific protein from a cell lysate.

Molecules have an ________ bond when they are held together by charge attraction.

ionic

Positively charged amino acids (3)

lysine (K), arginine (R), & histidine (H)

kinase.

Protein that transfers a phosphate group from ATP to an amino acid on another protein.

The release of hydrogen ions in a solution would result in _____________.

Lower pH

The solution consisting of the material released from disrupted or broken cells is called ______

lysate

_________________ antibodies are all identical and bind to a single epitope or antigen.

Monoclonal

Amino acids are to proteins as __________ are to carbohydrates.

monosaccharides

atoms that act as hydrogen bond donors

N, O, and F

A molecule that contains a five-carbon sugar, a phosphate group and a nitrogenous base is called a ____________

Nucleotide

Amino acids NOT found in alpha helices (single code)

P and G

Peptide Bond

Covalent bond formed between amino acids.

These are examples of hydrophobic amino acids.

Phe & Trp

phosphatase

Protein that removes a phosphate group from another protein

A molecule composed of a glycerol backbone bonded to two fatty acid chains and a small polar group is called a _____________.

Phospholipid

_________________ antibodies consist of a large collection of different antibodies what bind to different epitopes or antigens.

Polyclonal

In Western Blot experiments, the ____________ antibody is used to bind to the specific protein of interest.

primary

The tertiary structure of a protein is determined by its _____________.

Primary structure

Primary structure of proteins

the linear arrangement of amino acids in a protein; involves peptide bonds between amino acids.

Pulse

The window of time during which radioactively-labeled amino acids are fed to cells

Pulse-Chase

A method used to track a specific set of proteins through time

Quaternary structure of proteins

formed when two or more polypeptide chains specifically associate with one another to form a protein complex. It may or may not include disulfide bonds.

These amino acids can be phosphorylated

S, T, & Y

These amino acids have a hydroxyl (-OH) group (3, name + single code)

serine (S), threonin (T), and tyrosine (Y)

Methods used to separate proteins by size.

SDS-PAGE

In Western Blot experiments, the ____________ antibody is used to amplify the signal to detect the protein of interest.

secondary

Secondary structure of proteins

alpha helices and beta sheets; local structure formed via hydrogen bonds between amino acids; includes hydrogen bonds between atoms of the polypeptide chain NOT side chains

Sodium Dodecyl Sulfate (SDS)

strong detergent typically used to denature proteins and coat them with a negative charge before running them on a gel

Tertiary structure of proteins

overall polypeptide chain shape formed via non-covalent bonds between amino acids over long stretches of the chain. May include disulfide bonds

These amino acids can be modified by glycosylation of their hydroxyl R group.

Thr & Ser

van der Waals force

Noncovalent interaction between two molecules that are so close together that they can experience weak attractive forces bonding them together

Western Blot

Method used to stain a specific protein after its separation on a gel.

formation of covalent bonds is based on what principle?

atoms are most stable with a full outer electron shell

how to determine the number of covalent bonds?

number of electrons needed to fill the outer shell

atoms can be joined by bonds in which ____

> 1 pair of electrons are shared

how is the molecular shape determined?

type of bond; only single bonds can rotate relative to one another

describe the polarity of water (2)

• O-H bonds polarized due to O being partially negative, H partially positive

• asymmetric charge distribution

which elements will polar molecular typically have?

1+ electronegative atoms - O, N, S, P

what are nonpolar molecules?

molecules without electronegative atoms or polar bonds, consist of C & H

how to determine molecular reactivity?

presence of strongly polarized bonds

molecular reactivity for molecules without electronegative atoms (waxes & fats)

relatively inert

molecular reactivity for molecules with electronegative atoms

more reactive

what is ionization?

some atoms are so strongly electronegative that they can capture electrons from another atom during a chemical reaction

describe how table salt is ionized with Na and Cl (3)

1. single electron in Na outer shell migrates to electron-deficient chlorine atom

2. each atom becomes charged (ion; Cl- Na+)

3. atom forms crystal and is stable due to filled outer shell

when a different electron arrangement in atoms produces a highly reactive species, what is it called?

free radical

significance of noncovalent bonds?

govern interactions between molecules or different parts of a large biological molecule; usually weaker linkages but together can provide stability

types of noncovalent bonds?

ionic, hydrogen, van der waals forces, hydrophobic forces

how does the strength of an ionic bond differ in a crystal vs. in water vs. free vs. in cell vs. in protein core (5)?

crystal - strong

water - water prevents attraction between them

free - not important due to cells being mostly water

in cell - generally weak due to water

protein core - water excluded, so can be influential

what happens if hydrogen is bonded to an electronegative atom (O, N) (2)?

1. shared electron pair displaced towards electronegative atom so H is partially positive

2. H shared between two electronegative atoms

are hydrogen bonds hydrophobic or philic?

hydrophilic; enhance solubility in and interactions with water

what happens when a barely positively charged nucleus of H approaches an unshared pair of outer electrons of a second electronegative atom?

attractive (weak electrostatic) interaction (H-bond) occurs

where do H bonds mostly occur in and what impact do they have?

between most polar molecules; important in determining structure & properties of water, but also between polar groups & large biological molecules

what are van der waals interactions?

hydrophobic groups forming weak bonds with one another based on electrostatic interactions/due to slight perturbations of electron distributions

what is the symmetry of van der waals interactions look like (2)?

• electron distributions in nonpolar covalent bonds not always symmetric & vary moment to moment

• electron density may be larger on one side of an atom or the other; transient charge asymmetries result in momentary charge separations (dipoles)

how strong are van der waals forces (2)?

single forces very weak & very sensitive to distance; molecules must be close together & complementary shapes of interaction portions allow close approach

interactions can induce separation in adjacent molecules & lead to additional attractive forces among nonpolar molecules

significance of van der waals interactions?

important biologically with antibody/viral antigen interactions

what are hydrophobic forces?

molecules with nonpolar covalent bonds lacking a charged region that can interact with poles of water molecules & are thus insoluble in water

what elements are proteins generally composed of?

CHON & usually S or P

what is the amino acid backbone like?

alpha-carbon between NH2 and COOH groups; all but glycine have symmetric centers so there are 2 stereoisomers D- & L-amino acids

what form of stereoisomer is in proteins?

L-amino acid

in an aqueous environment -COOH ionizes to ____ and -NH2 ionizes to ____

-COO-, NH2-

how are peptide bonds formed?

condensation reactions attaching alpha-carboxyl of 1 amino acid to alpha-amino group of another using the N and C-terminus

significance of r groups (2)?

determine inter- & intramolecular interactions that determine molecular structure & protein activities, respectively

give amino acids their variability

significance of having a common amino acid backbone?

allows polymerization to form by same reaction no matter which two amino acids are being joined

what are the four amino acid & r group categories?

acidic (negatively charged), basic (positively charged), uncharged polar, nonpolar side chains

what are acidic/negatively charged amino acids?

have r-groups that act as stronger organic acids; can form ionic bonds

characteristics of acidic/negatively charged amino acids (2)?

• almost always fully negatively charged in cells (at pH 7, cellular pH)

• can form ionic bonds due to charges

which amino acids are acidic/negatively charged (2, include three/single codes)?

aspartic acid (asp/D), glutamic acid (glu/E)

what are basic/positively charged amino acids?

have r groups that act as stronger organic bases

which amino acids are basic/positively charged (3, include three/single codes)?

lysine (lys/K), arginine (Arg/R), histidine (His/H)

what are characteristics of basic/positively charged amino acids (2)?

• can form ionic bonds due to charges

• lys/arg are almost fully positively charged in cells; his partially positively charged

what are some characteristics of uncharged polar amino acids?

• tend to be in hydrophilic areas of protein, outside surfaces exposed to cytoplasm due to polarity

• serine, threonine, tyrosine have a hydroxyl group that can be modified by addition of other groups

which amino acids are uncharged polar (5, include three/single codes)?

asparagine (asn/N), glutamine (gln/Q), serine (ser/S), threonine (Thr/T), tyrosine (Tyr/Y)

which amino acids are nonpolar side chains (10, include three/single codes)?

alanine (ala/A), valine (val/V), leucine (leu/L), isoleucine (ile/I), proline (pro/P), phenylalanine (phe/F), methionine (met/M), tryptophan (tryp/W), glycine (gly/G), cysteine (cys/C)

what are some characteristics of nonpolar side chains (1)?

tend to reside in hydrophobic areas of the protein (ex: inner portion of a globular protein or the transmembrane domain of an integral membrane protein) due to nonpolarity

what is so special about cysteine?

r has reactive SH that can form disulfide bridges with other cysteines often at some distance away or in another chain. this stabilizes proteins, especially outside cells where they get added chemical & physical stress

how are most soluble proteins set up?

polar & charged residues on molecule surface