Amino acids chemical structure

AMINO ACIDS ARE ORGANIC COMPOUNDS MADE OUT OF CARBON HYDROGEN OXYGEN NITROGEN SULPHUR (IN A FEW CASES)

AMINO ACIDS CHEMICAL STRUCTURE

The basic structure of an amino acid molecule consists of

carbon atom that is bonded to an amino group (-NH2), carboxyl group (-COOH), hydrogen atom (H) R group or the side chain differs from one amino acid to another R group can vary widely and is responsible for the differences in the chemical properties

AMINO ACIDS CHEMICAL STRUCTURE

referred to as a-amino acids because the amino group is bonded to the a-carbon atom The name, amino acid, comes from the amino group and the acid group which are the most reactive parts of the molecule.

AMINO ACIDS CHEMICAL STRUCTURE

R H2NaC COOH H

R H2NaC H COOH AMINO ACID

CH3 H2 NaC H COOH IN ALANINE R GROUP IS CH3

OPTICAL PROPERTIES CHIRALITY

In chemistry, chirality refers to how a molecule's components are arranged, in space, about a central atom (or atoms). Chiral molecules lack symmetry and therefore cannot be superimposed on their mirror images

AMINO ACID CHIRALITY

Stereoisomerization Amino acids (except glycine) have a chiral carbon atom adjacent to the carboxyl group (COO-).

This chiral center allows for stereoisomerism. For example, take a look at the stereo representations and Fischer projection formulas of the enantiomers of serine

AMINO ACID CHIRALITY

AMINO ACID CHIRALITY

All amino acids found in proteins occur in the L-configuration about the chiral carbon atom. D-amino acids are not naturally found in proteins and are not involved in the metabolic pathways of eukaryotic organisms, D-amino acids are important in the structure and metabolism of bacteria. For example, Dglutamic acid and D-alanine are structural components of certain bacterial cell walls. D-amino acids are in polypeptide anitbiotics.

CHIRALITY

POLARIMETRY
a method to analyze chiral substances. The magnitude and direction of rotation of the plane of polarized light by a chiral compound is a specific physical property of the compound that can be used to characterize it Most biomolecules are chiral and hence rotate polarized light. Angle of rotation is depends on Concentration of the solution Path length of the solution Wave length of the light Chemical configuration of the substance

LIGHT WAVES

POLARIMETER

OPTICAL PROPERTIES LIGHT ABSORBANCE

The aromatic R-groups in amino acids absorb ultraviolet light with an absorbance maximum in the range of 280nm. The ability of proteins to absorb ultraviolet light is predominantly due to the presence of the tryptophan which strongly absorbs ultraviolet light. This group has alternative double bonds

EACH AMINO ACID DIFFERS FROM ONE ANOTHER, DUE TO THE NATURE OF R

Different amino acids

Classification Nature of the R group

TWENTY AMINO ACIDS APPEAR REGULARLY IN PROTEINS.
THESE AMINO ACIDS ARE CLASSIFIED ACCORDING TO THE NATURE OF THE R GROUP OF THE MOLECULE.

AMINO ACIDS WITH ALIPHATIC R-GROUPS

NON-AROMATIC AMINO ACIDS WITH HYDROXYL R-GROUPS

AMINO ACIDS WITH SULFUR-CONTAINING R-GROUPS

Acidic Amino Acids and their Amides

BASIC AMINO ACIDS

AMINO ACIDS WITH AROMATIC RINGS

IMINO ACIDS

PROLINE

AMINO ACID CLASSIFICATIONS There are two broad classes of amino acids based upon whether the R-group is

hydrophobic hydrophilic.

HYDROPHOBIC
The hydrophobic amino acids tend to repel the aqueous environment reside predominantly in the interior of proteins. does not ionize nor participate in the formation of H-bonds.

HYDROPHILC
The hydrophilic amino acids tend to interact with the aqueous environment,

are often involved in the formation of Hbonds and

are predominantly found on the exterior surfaces proteins or in the reactive centers of enzymes.

NON POLAR NEUTRAL

NON-POLAR SIDE CHAINS

Side chains which have pure hydrocarbon alkyl groups (alkane branches) or aromatic (benzene rings) are nonpolar. Examples include valine, alanine, leucine, isoleucine, phenylalanine.
The number of alkyl groups also influences the polarity. The more alkyl groups present, the more non-polar the amino acid will be. This effect makes valine more nonpolar than alanine leucine is more non-polar than valine.

CLASSIFICATION NON POLAR NEUTRAL

Neutral Side Chains both an amine and acid group which have been neutralized in the zwitterion, the amino acid is neutral unless there is an extra acid or base on the side chain. If neither is present then the whole amino acid is neutral.

CLASSIFICATION NON POLAR NEUTRAL

Neutral Side Chains Amino acids with an amide on the side chain do not produce basic solutions i.e. asparagine and glutamine. You need to look at the functional groups carefully because an amide starts out looking like an amine, but has the carbon double bond oxygen which changes the property. Amides are not basic. Even though tryptophan has an amine group as part of a five member ring, the electron withdrawing effects of the two ring systems do not allow nitrogen to act as a base by attracting hydrogen ions.

CLASSIFICATION NON POLAR NEUTRAL

CLASSIFICATION NON POLAR NEUTRAL

BASIC SIDE CHAINS

If the side chain contains an amine functional group, the amino acid produces a basic solution because the extra amine group is not neutralized by the acid group. Amino acids which have basic side chains include: lysine, arginine, and histidine. Amino acids with an amide on the side chain do not produce basic solutions i.e. asparagine and glutamine.

POLAR POSITIVE

POLAR SIDE CHAINS:

Side chains which have various functional groups such as acids, amides, alcohols, and amines will impart a more polar character to the amino acid. The ranking of polarity will depend on the relative ranking of polarity for various functional groups as determined in functional groups. In addition, the number of carbon-hydrogens in the alkane or aromatic portion of the side chain should be considered along with the functional group.

POLAR SIDE CHAINS:

Example: Aspartic acid is more polar than serine because an acid functional group is more polar than an alcohol group. Example: Serine is more polar than threonine since threonine has one more methyl group than serine. The methyl group gives a little more non-polar character to threonine. Example: Serine is more polar than tyrosine, since tyrosine has the hydrocarbon benzene ring.

ACIDIC SIDE CHAINS:

If the side chain contains an acid functional group, the whole amino acid produces an acidic solution. Normally, an amino acid produces a nearly neutral solution since the acid group and the basic amine group on the root amino acid neutralize each other in the zwitterion. If the amino acid structure contains two acid groups and one amine group, there is a net acid producing effect. The two acidic amino acids are aspartic and glutamic.

POLAR NEGATIVE

POLAR NEGATIVE

SWITTERION FORMATION

R R R H3N + C COOH H3N+ C COO - H2N C COO H H H

Cation (+) acidic Zwitterion (0) iso-electric pH Anion (-) alkaline

. PH DEPENDENCY

Since amino acids, as well as peptides and proteins, incorporate both acidic and basic functional groups, the predominant molecular species present in an aqueous solution will depend on the pH of the solution. In order to determine the nature of the molecular and ionic species that are present in aqueous solutions at different pH's, we make use of the Henderson-Hasselbach Equation,

PKA REPRESENTS THE ACIDITY OF A SPECIFIC CONJUGATE ACID FUNCTION (HA). WHEN THE PH OF THE SOLUTION EQUALS PKA, THE CONCENTRATIONS OF HA AND A(-) MUST BE EQUAL (LOG 1 = 0).

THE ISOELECTRIC POINT

the isoelectric point, pI, is the pH of an aqueous solution of an amino acid (or peptide) at which the molecules on average have no net charge. In other words, the positively charged groups are exactly balanced by the negatively charged groups. For simple amino acids such as alanine, the pI is an average of the pKa's of the carboxyl (2.34) and ammonium (9.69) groups. Thus, the pI for alanine is calculated to be: (2.34 + 9.69)/2 = 6.02, the experimentally determined value.

ALANINE TITRATION CURVE

AMINO ACID TITRATION CURVE GLUTAMIC ACID

ABBREVIATIONS AND DEFINITIONS

pK1: The pK of the carboxyl group of the amino acid. pK2: The pK of the amino group of the amino acid.

pKR: The pK of the side arm R of the amino acid. Only the specified AA's have ionizable side chains.

NINHYDRIN REACTION
specific

for amino acids a bluish purple coloured complex is formed with Ninhydrin. widely used in paper and ionexchange chromatography to visualize the separated amino acids.

THE NINHYDRIN REACTION TRIKETOHYDRINDENE HYDRATE

This Rpurple colored imino derivative, which provides as a useful color test for these amino acids, most of which are colorless

THE NINHYDRIN REACTION

CHROMATOGRAPHY
The ratio of the distance a compound moves from the baseline to the distance of the solvent front from the baseline is defined as the retardation (or retention) factor Rf. Different amino acids usually have different Rf's under suitable conditions.

CHROMATOGRAPHY

retardation (or retention) factor Rf

COMPOSITION OF PROTEINS

There are 20 -amino acids that are relevant to the make-up of mammalian proteins Several other amino acids are found in the body free or in combined states (i.e. not associated with peptides or proteins). These non-protein associated amino acids perform specialized functions. Several of the amino acids found in proteins also serve functions distinct from the formation of peptides and proteins, e.g., tyrosine in the formation of thyroid hormones. glutamate acting as a neurotransmitter

CLASSIFICATION OF AMINO ACID ESSENTIAL AND NON-ESSENTIAL

Essential Amino acids

Non-essential Amino Acids

1. Histidine 2. Isoleucine 3. Leucine 4. Lysine 5. Methionine 6. Phenylalanine 7. Threonine 8. Tryptophan 9. Valine

1. Alanine 2. Arginine* 3. Aspartic acid 4. Cysteine*

5. Cystine
6. Glutamic acid 7. Glutamine* 8. Glycine 9. Proline 10. Serine 11. Tyrosine

THE PEPTIDE BOND

Peptide bond formation is a condensation reaction leading to the polymerization of amino acids into peptides and proteins. Peptides are small consisting of few amino acids. A number of hormones and neurotransmitters are peptides. Additionally, several antibiotics and antitumor agents are peptides. Proteins are polypeptides of greatly divergent length.

THE PEPTIDE BOND

The simplest peptide, a dipeptide, contains a single peptide bond formed by the condensation of the carboxyl group of one amino acid with the amino group of the second with the concomitant elimination of water. The presence of the carbonyl group in a peptide bond allows electron resonance stabilization to occur such that the peptide bond exhibits rigidity not unlike the typical C=C double bond. The peptide bond is, therefore, said to have partial double-bond character.

R1 H2N C COOH H

-H2O

R2 H N C COOH H H

H2N C CO H

R1

R2 HN C COOH H

CONDENSATION OF TWO AMINO ACIDS MOLECULESTO FORM A DIPEPTIDE.

PEPTIDE BOND FORMATION

OLIGOPEPTIDES
NH2 CC - NH COOH COOH NH2 NH CH2 (CH2)2 (CH2)4 (CH2)3 H2N CCONHCCON CCONHCCOOH H H H H Aspartic glutamic lysine Arginine
**

BIURET REACTION

Peptides and proteins (long-chain polypeptides) react with Cu2+ in alkalinity to create a blue-violet colored chelate complex with an absorbance maximum at 540 nm. The peptide must have three peptide bonds at least.

BIURET REACTION

WITH AMINO ACIDS

Amino acids and Cu2+ ions form a blue complex.

THE BIURET TEST

used for the quantitative photometrical determination of total protein concentration. The intensity of the color produced in the biuret reaction is proportional to the number of peptide bonds participating in the reaction.

KJELDAHL METHOD FOR PROTEINS

The method consists of heating a substance with sulfuric acid, which decomposes the organic substance by oxidation to liberate the reduced nitrogen as ammonium sulfate. In this step potassium sulfate is added in order to increase the boiling point of the medium (from 337F to 373F / 169C to 189C). Chemical decomposition of the sample is complete when the medium has become clear and colorless (initially very dark).

KJELDAHL METHOD

The solution is then distilled with sodium hydroxide (added in small quantities) which converts the ammonium salt to ammonia. The amount of ammonia present (hence the amount of nitrogen present in the sample) is determined by back titration. The end of the condenser is dipped into a solution of boric acid. The ammonia reacts with the acid and the remainder of the acid is then titrated with a sodium carbonate solution with a methyl orange pH indicator.

Degradation: Protein + H2SO4 (NH4)2SO4(aq) + CO2(g) + SO2(g) + H2O(g) Liberation of ammonia: (NH4)2SO4(aq) + 2NaOH Na2SO4(aq) + 2H2O(l) + 2NH3(g) Capture of ammonia: B(OH)3 + H2O + NH3 NH4+ + B(OH)4 Back-titration: B(OH)3 + H2O + Na2CO3 NaHCO3(aq) + NaB(OH)4(aq) + CO2(g) + H2O

Nowadays, the Kjeldahl method is largely automated and makes use of specific catalysts (mercury oxide or copper sulfate) to speed up the decomposition.

KJELDAHL METHOD

The interferecnces of Kjeldhahl method Nitrogen containing substances such as Melamine and urea

Melamine

THE STRUCTURE OF amino acids. Proteins are polymers of PROTEINS

acid side chains has significant influence on the topography of the protein. The bonds between amino acid side chains generate a complex protein structure, which is considered in four stages: primary, secondary, tertiary, and quaternary. Four orders or levels of proteins structure

THE BACKBONE OF THE AMINO-ACID SEQUENCE LEU-ALA-GLU IS SHOWN IN PURPLE, AND THE SIDE CHAINS ARE SHOWN IN GREEN

PRIMARY STRUCTURE
Primary Structure refers to the linear sequence of amino acids that make up the polypeptide chain. This sequence is determined by the genetic code, the sequence of nucleotide bases in the DNA. The bond between two amino acids is a peptide bond. The sequence of amino acids determines the positioning of the different R groups relative to each other. This positioning therefore determines the way that the protein folds and the final structure of the molecule

PRIMARY PROTEIN STRUCTURE SEQUENCE OF CHAIN OF AMINO ACIDS

N TERMINAL C TERMINAL

THE SECONDARY STRUCTURE

refers to the shape the protein is pulled into by hydrogen bonds that form between the side chains of the amino acids. The secondary structure of protein molecules refers to the formation of a regular pattern of twists or kinks of the polypeptide chain. The regularity is due to hydrogen bonds forming between the atoms of the amino acid backbone of the polypeptide chain.

THE SECONDARY STRUCTURE

There are three common shapes formed: a -helix, b -pleated sheet, triple helix. Eg collagen All three shapes are very regular and exist as a result of hydrogen bonds between side chains that occur at regular intervals along the primary structure.

ANOTHER COMMON REPRESENTATION FOR PROTEINS AND PEPTIDES IS THE RIBBON, WHICH TRACES THE BACKBONE OF A PROTEIN OR PEPTIDE. THIS REPRESENTATION DOES NOT INCLUDE THE ATOMS IN THE SIDE CHAINS OF THE RESIDUES AND IS OFTEN USED TO REPRESENT THE THREE DIMENSIONAL STRUCTURE NOTICE THE BUNDLE OF HELICAL, OR COILED, PARTS OF THE BACKBONE

alpha-helix in a peptide chain of ferritin. The helical shape is held by hydrogen bonds between the -NH and CO functional groups in the backbone. the ribbon representation (showing only the trace of the backbone) is superimposed on a ball-and-stick representation, in which the non-hydrogen atoms are shown as spheres and the bonds are shown as sticks

HYDROGEN BONDING

Polypeptides contain numerous proton donors and acceptors both in their backbone and in the R-groups of the amino acids. The environment in which proteins are found also contains the ample H-bond donors and acceptors of the water molecule. H-bonding, therefore, occurs not only within and between polypeptide chains but with the surrounding aqueous medium.

HYDROGEN BOND

HYDROGEN BOND

IN AN -HELIX AND A -SHEET, ALL OF THE POSSIBLE HBONDS INVOLVING THE PEPTIDE BOND'S DONOR AND ACCEPTOR GROUPS ARE FORMED. THIS STABILIZES THE STRUCTURE.

IN AN -HELIX , THE R-GROUPS POINT OUTWARD FROM THE LONG AXIS OF THE HELIX

SECONDARY PROTEIN STRUCTURE SEQUENCE OF AMINO ACIDS ARE LINKED BY HYDROGEN BONDS

SECONDARY STRUCTURE

THE TERTIARY STRUCTURE

Tertiary structure refers to the three dimensional globular structure formed by bending and twisting of the polypeptide chain. This process often means that the linear sequence of amino acids is folded into a compact globular structure. The folding of the polypeptide chain is stabilized by multiple weak, noncovalent interactions. These interactions include:

THE TERTIARY STRUCTURE

These interactions include:

Hydrogen bonds that form when a Hydrogen atom is shared by two other atoms. Electrostatic interactions that occur between charged amino acid side chains. Electrostatic interactions are attractions between positive and negative sites on macromolecules. Hydrophobic interactions: During folding of the polypeptide chain, amino acids with a polar (water soluble) side chain are often found on the surface of the molecule while amino acids with non polar (water insoluble) side chain are buried in the interior. This means that the folded protein is soluble in water or aqueous solutions.

THE TERTIARY STRUCTURE

Covalent bonds may also contribute to tertiary structure. The amino acid, cysteine, has an SH group as part of its R group and therefore, the disulfide bond (S-S ) can form with an adjacent cysteine. For example, insulin has two polypeptide chains that are joined by two disulfide bonds.

If two cysteine side chains end up next to each other because of folding in the peptide chain, they can react to form a sulphur bridge. This is another covalent link. sometimes this link can be considered as part of the primary structure of the protein.

DISULFIDE BOND

DISULFIDE BOND FORMATION

Sulfur groups on cysteines may undergo oxidation to form disulfide bonds that are not normally present. Extra disulfide bonds can form when proteins are removed from their normal environment. Reducing agents such as dithiothreitol or mercaptoethanol are often added to prevent undesirable disulfiate bond formation

IONIC INTERACTIONS

Some amino acids (such as aspartic acid and glutamic acid) contain an extra -COOH group. Some amino acids (such as lysine) contain an extra -NH2 group. can transfer a hydrogen ion from the -COOH to the -NH2 group to form zwitterions just as in simple amino acids. can obviously get an ionic bond between the negative and the positive group if the chains folded in such a way that they were close to each other.

IONIC BONDS

VAN DER WAALS DISPERSION FORCES

Several amino acids have quite large hydrocarbon groups in their side chains. A few examples are shown below.

VAN DER WAALS DISPERSION FORCES

Temporary

fluctuating dipoles in one of these groups could induce opposite dipoles in another group on a nearby folded chain. The dispersion forces set up would be enough to hold the folded structure together

TERTIARY PROTEIN STRUCTURE CERTAIN ATTRACTIONS ARE PRESENT BETWEEN ALPHA HELICES AND PLEATED SHEETS

TERTIARY STRUCTURE

IN -SHEETS, THE H-BONDS BETWEEN THE PEPTIDE BOND DONOR AND ACCEPTOR GROUPS ARE MADE BETWEEN EITHER PARALLEL OR ANTI-PARALLEL POLYPEPTIDE CHAINS. THESE ARE ANTI-PARALLEL -SHEETS THE R-GROUPS POINT OUT OF AND INTO THE PLANE OF THE SHEET.

THE QUATERNARY STRUCTURE

some proteins contain more than one polypeptide chain, adding an additional level of structural organization: the association of the polypeptide chains. Each polypeptide chain in the protein is called a subunit. The subunits can be the same polypeptide chain or different ones. eg

THE QUATERNARY STRUCTURE

For example, the enzyme galactosidase is a tetramer, meaning that it is composed of four subunits, and the subunits are identical - each polypeptide chain has the same sequence of amino acids.

QUATERNARY STRUCTURE PROTEIN CONSISTING OF MORE THAN ONE AMINO ACID CHAIN

QUATERNARY STRUCTURE

PROLINE PEPTIDE BONDS ARE FOUND IN THE CIS CONFIGURATION ~100 TIMES AS OFTEN AS THOSE BETWEEN OTHER AMINO ACIDS. THERE IS NO H-BOND DONOR IN THE PROLINE PEPTIDE BOND AND THE PRESENCE OF PROLINE LEADS TO A BEND OR KINK IN THE POLYPEPTIDE CHAIN.

HAEMOGLOBIN

comprised of four polypeptide subunits, two with alpha helix secondary structure and two with beta pleated sheet form. All four components carry a heme group that can bind to oxygen, and all four components must be present to form haemoglobin. The shape of the haemaglobin affects its ability to carry oxygen, and travel freely throughout the circulatory system.

HAEMOGLOBIN

A condition that is a result of a malformed haemoglobin unit is sickle-cell anemia. position glutamic acid is replaced by a valine, (This is the result of a single base pair change at the DNA level.) an ionic cross-link is not formed resulting in a severe change of shape of the tertiary structure of the haemoglobin. The resultant shape is a crescent or sickle which reduces the oxygen carrying capacity of the red blood cell. Sickle cells are removed from circulation faster than normal cells resulting in anemia. They can also clump together causing blockages, pain and organ damage

HAEMOGLOBIN

HEMOGLOBIN ADULT HEMOGLOBIN IS A [(2):(2)] TETRAMERIC HEMEPROTEIN FOUND IN ERYTHROCYTES

CLASSIFICATION OF PROTEINS

Proteins are the most abundant organic macromolecules found in the cells and body fluids.
Molecular weight of proteins range from 68,000 (eg: haemoglobin) to 5.2 million (eg: b-lipoproteins). Proteins are of importance in all biological processes.

CLASSIFICATION OF PROTEINS
Classification system based on salient features of proteins * solubility * shape * function * physical properties * three dimensional structure

SOLUBILITY

The line of demarcation is not clear.

Classification of globulins Pseudoglobulins freely water soluble Euglobulins insoluble in salt free water

SHAPE
Based on axial ratio (ratio of length to breadth of the molecule. Globular proteins axial ratio less than 10. Fibrous proteins grater than 10 eg polypeptide chains or group of chains coiled in spiral or helix, cross linked by hydrogen and disulfide links. Eg. keratin, myosin

SHAPE OF PROTEINS
The shape of proteins is critical to their function, the shape is largely a result of the bonds that form between the side chains of amino acids that make the protein. primary purpose of the side chains in amino acids is to give proteins their shape, which dictates their function.

FUNCTIONS OF PROTEINS STRUCTURAL.

Body is composed of various structural proteins * Collagen * mucoproteins in connective tissues

FUNCTIONS OF PROTEINS

AS HORMONES

Some hormones are structurally proteins eg * thyroid stimulating hormones (TSH) * growth hormones (GH)

FUNCTIONS OF PROTEINS
Nutrition Metabolism of proteins released amino acids to the pool. used for synthesis of new proteins / nitrogenous compounds. deaminated to give CO2 and water (catabolize).

FUNCTIONS OF PROTEINS
Transport of small molecules

Small molecules are transported in the plasma combined with proteins, specially albumin.. Hormone, cortisol combines with protein, transcortin Iron transferrin nucleic acid nucleoproteins pigments such as Bilirubin conjugate with albumin thyroxin hormone combines with Thyroxin Binding Globulin (TBG) Toxic substances covert non toxic by binding to proteins

FUNCTIONS OF PROTEINS
Control of body water distribution Maintenance of oncotic pressure Albumin, mainly acts to maintain the oncotic pressure in body fluids prevent, accumulation of fluids in the body tissues to prevent oedema. Oncotic pressure osmotic pressure exerted due to the presence of albumin

Functions of proteins

HOST DEFENSE MECHANISM FROM FOREIGN ANTIGENS

ON EXPOSURE TO ANTIGENS, SUCH AS VIRUS AND BACTERIA, BODY SYNTHESIZES ANTIBODIES WHICH ARE IMMUNOGLOBULINS

Functions of proteins

BLOOD COAGULATION

MANY OF THE COAGULATION FACTORS ARE PROTEINS.

RECEPTORS
glycoprotein structures * * Cytoplasm (oestriol receptors) surface of the cell membranes (insulin receptors).

important in the action of hormones

Functions of proteins Catalytic action

ENZYMES ARE PROTEINS, SMALL AMOUNTS SPEEDS UP THE RATE OF A BIOLOGICAL REACTION, WITHOUT BEING USED UP.

FUNCTIONS OF PROTEINS
BUFFERING ACTION

Proteins act as buffers in the body, Proteins are negatively charged at body pH and act as bases accepting hydrogen ions.

CLASSIFICATION ACCORDING TO BIOLOGICAL FUNCTIONS

CLASSIFICATION ACCORDING TO BIOLOGICAL FUNCTIONS

CLASSIFICATION PHYSICAL PROPERTIES

Distinguish closely related proteins. Eg lipoproteins LDL, VLDL or depending on apo lipoproteins present

CLASSIFICATION THREE DIMENSIONAL STRUCTURE

basis of whether or not possesses quaternary structure.

CLASSIFICATION OF PROTEINS SIMPLE & COMPLEX

All proteins are polypeptides

Simple proteins Conjugated proteins

(complex proteins)

SIMPLE PROTEINS
these on hydrolysis give only amino acids Hydrolysis is the chemical reaction, in which proteins reacts with water to produce amino acids

CONJUGATED PROTEINS
Some proteins are conjugated with nonprotein molecules. conjugated with

Carbohydrate glycoproteins lipids lipoproteins nucleic acid nucleoproteins pigments such as Bilirubin conjugate with albumin

CLASSIFICATION OF PROTEINS ACCORDING TO SHAPE AND SOLUBILITY

Proteins can be broadly classified into three groups, based on their shape and solubility.
Fibrous proteins: these proteins have a rod like structure. They are not soluble in water collagen is an example of a fibrous protein. Globular proteins: these proteins more or less spherical in nature. Due to their distribution of amino acids (hydrophobic inside, hydrophillic outside) they are very soluble in aqueous solution. Myoglobin is an example of a globular protein.

CLASSIFICATION OF PROTEINS ACCORDING TO SHAPE AND SOLUBILITY

Membrane proteins: these are protein which are in association with lipid membranes. Those membrane proteins that are embedded in the lipid bilayer have extensive hydrophobic amino acids that interact with the non-polar environment of the bilayer interior. Membrane proteins are not soluble in aqueous solution. Rhodopsin is an example of a membrane protein. Note that rhodopsin is an integral membrane protein and is embedded in the bilayer.

FUNCTIONAL SIGNIFICANCE OF AMINO ACID R-GROUPS In solution it is the nature of the amino acid R-groups that dictate structure-function relationships of peptides and proteins. The hydrophobic amino acids will generally be encountered in the interior of proteins shielded from direct contact with water. Conversely, the hydrophilic amino acids are generally found on the exterior of proteins as well as in the active centers of enzymatically active proteins. Indeed, it is the very nature of certain amino acid R-groups that allow enzyme reactions to occur.

FUNCTIONAL SIGNIFICANCE OF AMINO ACID RGROUPS

The imidazole ring of histidine allows it to act as either a proton donor or acceptor at physiological pH. Hence, it is frequently found in the reactive center of enzymes. ability of histidines in hemoglobin to buffer the H+ ions from carbonic acid ionization in red blood cells. It is this property of hemoglobin that allows it to exchange O2 and CO2 at the tissues or lungs, respectively.

DENATURATION OF PROTEINS
The natural or native structures of proteins may be altered, and their biological activity changed or destroyed by treatment that does not disrupt the primary structure. This denaturation is often done deliberately in the course of separating and purifying proteins. For example, many soluble globular proteins precipitate if the pH of the solution is set at the pI of the protein.

DENATURATION OF PROTEINS

Also, addition of trichloroacetic acid or the bisamide urea (NH2CONH2) is commonly used to effect protein precipitation. Following denaturation, some proteins will return to their native structures under proper conditions; but extreme conditions, such as strong heating, usually cause irreversible change.

DENATURATION. THE POLYPEPTIDE CHAIN HAS LOST ITS HIGHER ORDER STRUCTURE AND IS NOW A RANDOM COIL.

DENATURATION OF PROTEINS
Some treatments known to denature proteins are. Denaturing Action-Mechanism of Operation Heat - hydrogen bonds are broken by increased translational and vibrational energy.(coagulation of egg white albumin on frying.)

DENATURATION OF PROTEINS

Urea Solution competition for hydrogen bonds. (precipitation of soluble proteins.) Some Organic Solvents (e.g. ethanol & acetone)change in dielectric constant and hydration of ionic groups. (disinfectant action and precipitation of protein.) Agitation - shearing of hydrogen bonds. (beating egg white albumin into a meringue.)

PROTEOLYSIS

Proteins can also be broken apart by enzymes, called proteases, that digest the covalent peptide bonds between amino acids that are responsible for the primary structure. This process is called proteolysis and is irreversible. Cells contain proteases that are found in lysosomes, membrane bound organelles inside the cell. When cells are disrupted, lysosomes break and release these proteases, which can damage the other proteins in the cell.

PROTEOLYSIS

In the laboratory, it is therefore necessary to minimize the activities of cellular proteases to protect proteins from proteolysis. Methods used to minimize proteolysis include working at lower temperatures (4C), and adding chemicals that inhibit protease activity.

ADSORB TO SURFACE

Proteins readily adsorb (stick to) surfaces, thereby reducing their available activity. To prevent significant loss, do not store dilute solutions of proteins for prolonged periods of time. Always dilute them right before use.

hemoglobin electrophoresis

LANES 1 AND 5 ARE HEMOGLOBIN STANDARDS. LANE 2 IS A NORMAL ADULT. LANE 3 IS A NORMAL NEONATE. LANE 4 IS A HOMOZYGOUS HBS INDIVIDUAL. LANES 6 AND 8 ARE HETEROZYGOUS SICKLE INDIVIDUALS. LANE 7 IS A SC DISEASE INDIVIDUAL.

SYLLABUS

Amino acids and protein Chemical nature of amino acids, classification, optical properties, functional properties of Rgroups, acid -base properties of proteins, characteristics of the peptide bond, classification of proteins-primary, secondary, tertiary and quaternary structure of proteins, forces controlling the protein structures, denaturation of protein.Practical (3hrs)

THE END

THANK YOU!

THE QUATERNARY STRUCTURE

Hemoglobin,

the oxygen carrying protein in the blood, is also a tetramer but it is composed of two polypeptide chains of one type (141 amino acids) and two of a different type (146 amino acids). In chemical shorthand, this is referred to as a22 . For some proteins, quaternary structure is required for full activity (function) of the protein

FORCES CONTROLLING PROTEIN STRUCTURE

Hydrophobic Forces Proteins are composed of amino acids that contain either hydrophilic or hydrophobic R-groups. It is the nature of the interaction of the different R-groups with the aqueous environment that plays the major role in shaping protein structure. The spontaneous folded state of globular proteins is a reflection of a balance between the opposing energetics of H-bonding between hydrophilic R-groups and the aqueous environment and the repulsion from the aqueous environment by the hydrophobic R-groups. The hydrophobicity of certain amino acid R-groups tends to drive them away from the exterior of proteins and into the interior. This driving force restricts the available conformations into which a protein may fold.

FORCES CONTROLLING PROTEIN STRUCTURE

Electrostatic Forces Electrostatic forces are mainly of three types; chargecharge, charge-dipole and dipole-dipole. Typical chargecharge interactions that favor protein folding are those between oppositely charged R-groups such as K or R and D or E. A substantial component of the energy involved in protein folding is charge-dipole interactions. This refers to the interaction of ionized R-groups of amino acids with the dipole of the water molecule. The slight dipole moment that exist in the polar R-groups of amino acid also influences their interaction with water. It is, therefore, understandable that the majority of the amino acids found on the exterior surfaces of globular proteins contain charged or polar Rgroups.

FORCES CONTROLLING PROTEIN STRUCTURE

VAN DER WAALS FORCES

There are both attractive and repulsive van der Waals forces that control protein folding. Attractive van der Waals forces involve the interactions among induced dipoles that arise from fluctuations in the charge densities that occur between adjacent uncharged nonbonded atoms. Repulsive van der Waals forces involve the interactions that occur when uncharged non-bonded atoms come very close together but do not induce dipoles. The repulsion is the result of the electronelectron repulsion that occurs as two clouds of electrons begin to overlap. Although van der Waals forces are extremely weak, relative to other forces governing conformation, it is the huge number of such interactions that occur in large protein molecules that make them significant to the folding of proteins.

The key factor that determines the specific details of polypeptide folding is the interaction between R-groups, not only with one another but with water and dissolved ions. In a water soluble polypeptide, most but not necessarily all of the surface R-groups will by hydrophilic. Hydrophobic groups will tend to be buried in the polypeptide's interior. Some polypeptides are inserted into membranes. In these polypeptides, hydrophobic R-groups on the surface of the folded polypeptide will interact with the hydrophobic interior of the lipid bilayer.

POST TRANSLATIONAL MODIFICATIONS

IN ADDITION there are other amino acids in proteins: Naturally occuring chemical modifications of the 20 genetically encoded amino acids, eg. Hydroxylation of proline and lysine Phosphorylation of Serine & Tyrosine R-group methylation of lysine, histidine and arginine. R-group acetylation of lysine. N-terminal methylation of alanine. N-terminal acetylation of serine. N-terminal formylation of methionine. Addition of carbohydrates (at Asparagine and Serine). Addition of Lipids (at Cysteine, N-terminal glycine and C-terminal via carbohydrate linker group) Covalent attachment of cofactors (eg. pyridoxal-5-phosphate attached at lysine; hemes attached at cysteines).

FUNCTIONAL SIGNIFICANCE OF AMINO ACID RGROUPS

In solution it is the nature of the amino acid R-groups that dictate structure-function relationships of peptides and proteins. The hydrophobic amino acids will generally be encountered in the interior of proteins shielded from direct contact with water. Conversely, the hydrophilic amino acids are generally found on the exterior of proteins as well as in the active centers of enzymatically active proteins. Indeed, it is the very nature of certain amino acid R-groups that allow enzyme reactions to occur. The imidazole ring of histidine allows it to act as either a proton donor or acceptor at physiological pH. Hence, it is frequently found in the reactive center of enzymes. Equally important is the ability of histidines in hemoglobin to buffer the H+ ions from carbonic acid ionization in red blood cells. It is this property of hemoglobin that allows it to exchange O2 and CO2 at the tissues or lungs, respectively.

FUNCTIONAL SIGNIFICANCE OF AMINO ACID R-GROUPS

The primary alcohol of serine and threonine as well as the thiol (SH) of cysteine allow these amino acids to act as nucleophiles during enzymatic catalysis. Additionally, the thiol of cysteine is able to form a disulfide bond with other cysteines: Cysteine-SH + HS-Cysteine <> Cysteine-S-S-Cysteine This simple disulfide is identified as cystine. The formation of disulfide bonds between cysteines present within proteins is important to the formation of active structural domains in a large number of proteins. Disulfide bonding between cysteines in different polypeptide chains of oligomeric proteins plays a crucial role in ordering the structure of complex proteins, e.g. the insulin receptor. back to the top