Protein can be classified according to solubility, shape or the presence of nonprotein groups, etc.

For example:1. Solubility, two major families are the globular and

fibrous protein. The globular proteins are compact, are roughly spherical or ovoid in shape, and have axial ratios of not over 3 (the ratio of their shortes to longest dimention).

2. Composition. For example: glycoproteins, lipoproteins, metaloproteins (that incorporate a metal ion such as many enzyme do) etc.

3. Biologycal functions: enzymes, hormones, neurotransmitters, toxin, contractile muscle (myosin and actin), storage protein (casein, ovalbumin and ferritin), transfort protein (hemoglobin), structural proteins (collagen, elastine, and protein cell membranes) and protective proteins.

Amino Acid and Protein 1

Globular ProteinsMyoglobin, a monomeric protein of red

muscle, stores oxygen.Hemoglobin, a tetramic (22) protein of

eritrocytes, transport O2 to the tissue and return CO2 and rptons to the lung. Despite different primary structures, the secondary-tertiary structure of subunits of hemoglobin (Hb S), Val replaces the 6 Glu of Hb A. The genetic defect has known as thalassemia result from theh partial or total absence of one or more or chains of hemoglobin.

Amino Acid and Protein 2

Amino Acids and the Primary Stucture of Proteins

Prentice Hall c2002 Chapter 3 3

Important biological functions of proteins1. Enzymes, the biochemical catalysts2. Storage and transport of biochemical molecules3. Physical cell support and shape (tubulin, actin,

collagen)4. Mechanical movement (flagella, mitosis,

muscles)(continued)

Globular proteins

Prentice Hall c2002 Chapter 3 4

• Usually water soluble, compact, roughly spherical

• Hydrophobic interior, hydrophilic surface• Globular proteins include enzymes,carrier

and regulatory proteins

Fibrous proteins

Prentice Hall c2002 Chapter 3 5

• Provide mechanical support• Often assembled into large cables or threads• -Keratins: major components of hair and nails• Collagen: major component of tendons, skin,

bones and teeth

Amino Acid and Protein 6

MYOGLOBIN STRUCTURE

Amino Acid and Protein 7

Fibrous ProteinsCollagen is the most abundant of the fibrous

proteins that constitute more than 25% of the protein mass in the human body. These proteins in bone, teeth, tendons, skin, and soft connective tissue. Collagen forms a unique triple helix. Every third amino acid residue in collagen is a glycine residue. Collagen is also rich in proline and hydroxyproline, yielding a repetitive Gly-X-Y pattern in which Y generally is proline or hydroxyproline (Gly-X-Y-Gly-X-Y-Gly-X-Y-). Disease of collagen maturation include the vitamin C deficiency disease scurvy and Ehlers-Danlos syndrome.

Amino Acid and Protein 8

Amino Acid and Protein 9

Prions-ProteinHuman prionrelated protein, PrP, a glycoprotein

encoded on the short arm of chromosome 20, normally is monomeric and rich helix. Pathologic prion proteins, known as PrPc, is rich in sheet with many hydrophobic aminoacyl side chains. Prion disease are protein conformation diseases transmitted by altering the conformation, fatal neurogenerative diseases characterized by spongiform changes.

For example: Creutzfeld-Jacob disease in humans, scrapie in sheep, and bovine spongiform encephalopathy (mad cow disease) in cattle.

Amino Acid and Protein 10

ELECTROPHORESIS

Amino Acid and Protein 11

Analysis of Biomolecules

B. Amphoteric properties

Amino acids are amphoteric molecules ; that is, they have both basic and acidic groups

Monoamino-monocarboxylic acids exist in solution neutral pH are predominantly dipolar ions (or zwitter ion). In dipolar form of an amino acid, the amino group is protonated and positively charged (-NH3

+) and the carboxyl group is dissociated and negatively charged (-COO-)

Amino Acid and Protein 12continued

H O

R – C – C O–

NH2

Cationic form charge +1 pH IEP

Zwitter ion form charge 0 pH IEP

Anionic form charge -1 pH IEP

H O

R – C – C OH

NH3+

H O

R – C – C O–

NH3+

OH–

H+

OH–

H+

Basic ConceptsFundamental to electrophoretical separations is the fact that proteins areelectrically charges particles.The charges are derived from aminoacids with ionogenic side groups.

Amino Acid and Protein 13

Amino group

HR C NH2

C = OOH Acid group

ELECTROPHORESISElectrophoresis is a method for analysis

(this is separation or isolation) and characterization of biological polymers.

Here is the use of electrophoresis for the separation of proteins.

The sample containing the proteins to be separated is placed in an electric field which forces the electrically charged proteins to move.

Amino Acid and Protein 14

ELECTROPHORESIS

The movement of molecules (in an electric field) influenced by the

Size Charge Shape Chemical composition

of the molecule

Amino Acid and Protein 15

SEPARATION OF PROTEINSThe separation is normally performed

not in free solution but in a supporting gel medium.

The gel can either act as an ”inert” support for the electrophoresis buffer or actively participate in the separation by interacting with the proteins

In the latter case the protein-gel interaction is the actual separation factor while the electrical field merely makes the proteins migrate through the gel

Amino Acid and Protein 16

Methods of ElectrophoresisThe major difference between methods is the

type of support medium, which can be either cellulose or thin gels

Amino Acid and Protein 17

1. Cellulose is used as a support medium for low-molecular weight biochemicals such as amino acids and carbohydrates

2. Polyacrylamide and agarose gels are widely used as support media for larger molecules.

Methods of electrophoresis divided into

1. Polyacrylamide Gel Electrophoresis (PAGE)2. Discontinuous Gel Electrophoresis3. Sodium Dodecyl Sulfate-PAGE (SDS-PAGE)4. Pulsed Field Gel Electrophoresis (PFGE)5. Isoelectric Focusing of Protein (IEF)6. Capillary Electrophoresis (CE)7. Immunoelectrophoresis (IE)

Amino Acid and Protein 18