Specific Proteins

Question 1

Separates protein according to their electrical charges (usually at pH 8.6)

Answer 1

Protein electrophoresis

Question 2

Major proteins in plasma that contribute to the electrophoretic pattern are:

Answer 2

Albumin
Alpha1 anti trypsin
Alpha2 macro globulin
Haptoglobin
Beta lipoprotein
Transferrin
Complement C3
Fibrinogen
Immunoglobulin

Question 3

Plays several roles, including maintaining oncotic pressure, transporting small molecules and promoting or inhibiting inflammatory reactions

Answer 3

Plasma proteins

Question 4

The major clinical of serum and urine protein electrophoresis is to screen for:

Answer 4

Monoclonal gammopathies

Question 5

Methods to quantitate and fractionate proteins:

Answer 5

Turbidimetry
Colorimetry
Absorption spectrophotometry
Dye binding
Column chromatography
Electrophoresis
Immunoassays

Question 6

Process of unfolding the protein (using heat, pH or chemical compounds)

Answer 6

Denaturation

Question 7

The final shape of protein, has the lowest free energy possible

Answer 7

Conformation

Question 8

Protein shape determined by amino acid sequence

Answer 8

Protein folding

Question 9

Consist of repeating sequence of amino acids

Answer 9

Polypeptide backbone

Question 10

Important numbers: Proteins

Answer 10

16% nitrogen which differentiates it from CHO and lipids
15% make up of the cell
20 amino acids linked by peptide bonds

Question 11

Simple protein: symmetrical, compactly folded polypeptide chains (albumin)

Answer 11

Globular

Question 12

Simple protein: elongated, asymmetrical polypeptide chains (troponin and collagen)

Answer 12

Fibrous

Question 13

Conjugated proteins: with a metal prosthetic group

Answer 13

Metalloproteins

Question 14

Conjugated proteins: lipid prosthetic group

Answer 14

Lipoprotein

Question 15

Conjugated proteins: with 10-40% CHO attachment ( haptoglobin)

Answer 15

Glycoproteins

Question 16

Conjugated proteins: with >40% CHO attached (mucin)

Answer 16

Mucoprotein

Question 17

Conjugated proteins: with DNA or RNA attached (chromatin)

Answer 17

Nucleoproteins

Question 18

The A.A are linked to each other through covalent peptide bonding in a specific sequence to form polypeptide chain

Answer 18

Primary Structure

Question 19

Hydrogen bonds in peptide backbone; forms alpha and beta pleated sheets

Answer 19

Secondary structure

Question 20

Non covalent interaction; coiled polypeptide chain folds upon itself to form a 3 dimensional structure

Answer 20

Tertiary structure

Question 21

Two or more folded polypeptide chains bind to each other through hydrogen bonds and electrostatic interactions to form a functional protein

Answer 21

Quaternary structure

Question 22

Separated proteins dissolved in an electrolyte solution by application of an electric current through u shaped quartz tube

Answer 22

Electrophoresis

Question 23

Separation of the protein fractions into discrete bands or zones due to introduction of filter paper as an anti convection support medium

Answer 23

Zonal electrophoresis

Question 24

The buffer flow which carries the proteins with mechanical flow, not by charge (agar)

Answer 24

Endosmosis/ electro-osmosis

Question 25

Principles of endosmosis/ electro-osmosis

Answer 25

Positively charged ion - free to move under electromotive force - flow of buffer towards the CATHODE Negatively charged ion - tends to move it towards the ANODE

Question 26

Determines the amount of current and the movement of the proteins for a fixed voltage

Answer 26

Ionic strength of the buffer

Question 27

Principles of Ionic strength of the buffer:

Answer 27

If low, more current is carried by the charged proteins If high, less current is carried by the proeins, shorter distance reached

Question 28

Support medium for ionic strength:

Answer 28

Cellulose acetate membrane Agarose gel Starch gel Polyacrylamide gel

Question 29

Devised to resolve albumin and globulins into 2 or more fractions that can be measured for protein content

Answer 29

Precipitation

Question 30

Globulins precipate through addition of:

Answer 30

Na sulfate Na sulfite ammonium sulfate Methanol

Question 31

Used extensively as it accentuates abnormalities in serum protein composition, which involves depression of albumin and elevation of more globulin fractions

Answer 31

A/G ratio

Question 32

Simplest and mildest of all chromatography techniques, separation is based on differences in size

Answer 32

Size exclusion chromatography / SEC

Question 33

Principles of SEC:

Answer 33

Size matters: Large proteins will elute first and the smallest ones will elute last All CHON species continuously move through gel filtration columns all at the same time but at different rates

Question 34

Based on the reversible interaction between a charged protein and an oppositely charged chromatography medium

Answer 34

Ion exchanged chromatography (IEC)

Question 35

Proteins are usually applied at basic pH 8.6 at which they may negatively charged or may have no net charge

Answer 35

Anion Exchange chromatography

Question 36

Principles of anion exchange chromatography:

Answer 36

Neutral CHON pass through an anion exchange column | Anionic proteins stick to the (+) charged column matrix

Question 37

Begins at aid pH, with proteins having positive charge and adhering to a negatively charged column matrix

Answer 37

Cation exchange chromatography

Question 38

Samples are applied at high salt and are eluted with low salt. The support medium interacts with proteins with hydrophobic nature

Answer 38

Hydrophobic chromatography

Question 39

Specific binding between a protein of interest and another protein that has been covalently linked to a solid support medium of column

Answer 39

Affinity chromatography

Question 40

The most common use of affinity chromatography is for:

Answer 40

Purification of recombinant proteins

Question 41

Based on the flow through a capillary tube that can be tailored to resolution of different molecules based on size, hydrophobicity or stereospecificity

Answer 41

Capillary electrophoresis

Question 42

The ultimate reference method for determining concentration of proteins

Answer 42

Nitrogen analysis

Question 43

Consists of acid digestion to release ammonium ions from nitrogen containing compounds

Answer 43

Kjeldahl technique ( the reference method)

Question 44

In which double iodides (K and Hg) forms a colored complex with ammonia in an alkali medium

Answer 44

Nesslerization

Question 45

Used to estimate specific gravity and by interference, protein content

Answer 45

Cooper sulfate solution

Question 46

Protein in solution absorbs ultraviolet light at 280 nm

Answer 46

Absorbance

Question 47

Detects turbidity produced by precipitation of a reagent antibody with its target protein in a serum sample

Answer 47

Nephelometry

Question 48

Measures major serum proteins in automated immunochemical analyzers

Answer 48

Nephelometry

Question 49

Immunologic method that detects proteins in lower concentration

Answer 49

radioimmunoassay (RIA) and ELISA

Question 50

Standard dyes for electrophoresis:

Answer 50

Coomassie brilliant blue Ponceau S Amido black

Question 51

Possible by virtue of the specific binding of albumin to certain dyes. Dyes bound to albumin absorb maximally at slightly different wavelengths

Answer 51

Quantification of albumin

Question 52

Dyes used for quantitation of albumin:

Answer 52

``` Bromphenol blue Methyl orange Hydroxybenzenebenzoic acid Bromcresol purple Bromcresol green ```

Question 53

Protein forms precipitate on the addition of trichoroacetic acid and sulfosalicylic acid but not specific to proteins

Answer 53

Turbidimetric methods

Question 54

Highly specific for proteins and peptides that uses biuret method in which copper salts in alkaline solution forms a purple complex with substances containing 2 or more peptide bonds

Answer 54

Colorimetric technique

Question 55

Oxidizes phenolic compounds so that a greater sensitivity can be obtained giving a deep blue color

Answer 55

Folin ciocalteu reagent

Question 56

Uses biuret method plus phenol reagent to enhanced color formation

Answer 56

Lowey assay