Amino acids

Jana NovotnáDept. of Biochemistry

• Amino acids are building blocks of proteins.

• 20 common AA - encoded by standard genetic code, construct proteins in all species .

• Primary structure determinates unique three-dimensional structure, function, binding sites of proteins for different interactions.

• AA are important intermediates in metabolism (porphyrins, purines, pyrimidines, creatin, urea etc).

• AA can have hormonal and catalytic function.

• Several genetic disorders are cause in amino acid metabolism errors (aminoaciduria - presence of amino acids in urine)

The basic structure of amino acids

Simple monoamino monocarboxyl a-amino acid

- diprotic acid - can yield protons when fully protonated

Binding interactions of amino acids

Electrostatic interactions

A disulfide bond

Dissociation of the side chains of amino acids

The Stereochemistry of Amino Acids

Chiral molecules existing in two forms

http://www.imb-jena.de/~rake/Bioinformatics_WEB/gifs/amino_acids_chiral.gif

The two stereoisomers of alanine

-a carbon is a chiral center

Two stereoisomers are called enantiomers.

The solid wedge-shaped bonds project out of the plane of paper, the dashed bonds behind it.

The horizontal bonds project out of the plane of paper, the vertical bonds behind.

Classification Based on Chemical Constitution

Small amino acids – Glycine, Alanine

Branched amino acids – Valine, Leucine, Isoleucine

Hydroxy amino acids (-OH group) – Serine, Threonine

Sulfur amino acids – Cysteine, Methionine

Aromatic amino acids – Phenylalanine, Tyrosine,

Tryptophan

Acidic amino acids and their derivatives –

Aspartate, Asparagine, Glutamate, Glutamine

Basic amino acids – Lysine, Arginine, Histidine

Imino acid - Proline

Essential and Nonessential Amino Acids

Arginine* Histidine* Isoleucine Leucine Valine

Lysine Methionine Threonine Phenylalanine Tryptophan

Alanine Asparagine Aspartate Glutamate Glutamine

Glycine Proline Serine Cysteine (from Met) Tyrosine (from Phe)

* Essential in children, not in adults

Uncommon amino acids found in proteins

Intermediates of biosynthesis of arginin and in urea cycle

Peptide Bond Formation

Proteins

The three-dimensional structure is determined by amino acid sequence.

The function depends on the structure.

An isolated protein exist in one or a small number of stable structural form.

The most important forces stabilizing the specific structure are noncovalent interactions.

How a sequence of amino acids in a polypeptide chain is translated into a discrete, three dimensional protein

structure?

The Peptide Bond Is Rigid and Planar

The peptide bond is a hybride between the resonance forms – the carbonyl oxygen has a partial negative charge and the amide nitrogen a partial positive charge, partial double form of peptide bond itself.

The N-Ca and Ca-C can rotate on angles f and j, resp., the peptide C-N bond is not free to rotate.

Take over from: D. L. Nelson, M. M. Cox :LEHNINGER. PRINCIPLES OF BIOCHEMISTRY Fifth edition

Knowledge of primary structure of protein is require for

understanding of :

the protein´s structure

the mechanism of protein action on molecular level

the interrelationship with other proteins in evolution

Sequencing of protein is an aids for :

the study of protein modification

the prediction of the similarity between two proteins

The determination of the primary structure of a protein requires a

purified protein.

The cloning of the genes for many proteins and the sequencing of

gene is a much faster method to obtain the amino acid sequence.

Primary Structure of Proteins

The primary structure of peptides and proteins refers to the linear number and order of the amino acids present.

the N-terminal end - on the left (the end bearing the residue with the free a-amino group)

the C-terminal end - on the right (the end with the

residue containing a free a-carboxyl group) .

Knowledge of primary structure of insulin aids in understanding its synthesis and action.

1. Pancreas produces single chain precursor – proinsulin

2. Proteolytic hydrolysis of 35 amino acid segment – C peptide

3. The remainder is active insulin(two polypeptide chains A and B) covalently joined by disulfide bonds

Amino acid identity in different animals:

Human, hors, rat, pig, sheep, chicken insulin have differences only in residues 8, 9, and 10 of the A chain and residue 30 of the chain B

Secondary structureThe second level of protein structure determined by attractive and repulsive forces among the amino acids in the chain. It is the specific geometric shape caused by intra-molecular and intermolecular hydrogen bonding of amide groups.Tertiary structureThree dimensional structure of polypeptide units (includes conformational relationships in space of side chains R of polypeptide chain).Quaternary structure Polypeptide subunits non-covalently interact and organize into multi-subunit protein (not all proteins have quaternary structure).

The folding of the primary structure into secondary, tertiary and quaternary structure appears to occur in most cases spontaneously.Cystein disulfide bonds are made after folding

HIGHER LEVELS OF PROTEIN ORGANIZATION

The a Helix

a-helix is a right-handed coiled conformation.

Every backbone N-H group of peptide bond donates a hydrogen bond to the backbone C=O group of the amino acid four residues earlier.

3.6 amino acid residues are per 360o turn.

The formation of the a-helix is spontaneous.

Protein Secondary Structure

b–Sheets

2 strands (segments) of polypeptide chains are stabilized by H-bonding between amide nitrogens and carbonyl carbons.

Polypeptide segments are aligned in parallel or anti-parallel direction to its neighboring chains.

b-structure gives plated sheet appearance – side chain groups are projected above and below the plane generated by the hydrogen-bonded polypeptide chains.

In parallel sheets adjacent peptide chains proceed in the same direction (i.e. the direction of N-terminal to C-terminal ends is the same).

In anti-parallel sheets adjacent chains are aligned in opposite directions.

The large number of hydrogen bonds maintain the structure in a stretched shape.

b–Sheets

The folding pattern of the secondary structural element into 3D conformation

a helical regions

b plated sheets

Forces that give rise to tertiary structureThe tertiary structure of a protein

Hydrophobic interaction

Protein Tertiary Structure

Examples of ,a b-folded domains in which b-structural strands form a b barrel in the centre of the domain

Examples of b-folded domains

Examples of the Tertiary Structure

Protein Quaternary Structure

Four subunits (two a and two b subunits) are associated in the quaternary structure

Hemoglobin

The arrangement of the protein subunit in the three-dimensional complex constitutes quaternary structure.

Forces Controlling Protein Structure

Hydrophobic interaction forces: Interaction inside polypeptide chains (amino acids contain either

hydrophilic or hydrophobic R-groups). Interaction between the different R-groups of amino acids in

polypeptide chains with the aqueous environment.

A non-polar residues dissolved in water induces in the water solvent a solvation shell in which water molecules are highly ordered.

Two non-polar groups in the solvation shell reduce surface area exposed to solvent and come very close come together.

Hydrogen bonds: Proton donors and acceptors within and between polypeptide

chains (backbone and the R-groups of the amino acids). H-bonding between polypeptide chains and surrounding aqueous

medium.

Electrostatic forces: Charge-charge interactions between oppositely charged R-

groups such as Lys or Arg (positively charged) and Asp or Glu (negatively charged).

Ionized R-groups of amino acids with the dipole of the water molecule.

van der Waals forces: Weak non-colvalent forces of great importance in protein

structure, the sum of the attractive or repulsive forces between molecules

Force is caused by the attraction between electron-rich regions of one molecule and electron-poor regions of another

Protein Denaturation and FoldingDenaturation is a loss of the three-

dimensional. The protein loss of it function.

Denaturation by heat has complex effect on the weak interactions (primarily by disrupting hydrogen bonds).

Extremes of pH alter the net charges on

the protein, causing electrostatic repulsion and the disruption of some hydrogen bonding.

Organic solvents and detergents act primarily by disrupting hydrophobic interactions

Renaturation is process in which protein regains its native structure

Take over from: D. L. Nelson, M. M. Cox :LEHNINGER. PRINCIPLES OF BIOCHEMISTRY Fifth edition

Some Proteins Undergo Assisted Folding

Not all proteins fold spontaneously and require molecular chaperons. Chaperons interact with partially or improperly folded polypeptides

Take over from: D. L. Nelson, M. M. Cox :LEHNINGER. PRINCIPLES OF BIOCHEMISTRY Fifth edition

Protein Misfolding

Amyloid fibre is an insoluble extracellular formation (amyloidoses). They arise from at least 18 inappropriately folded versions of proteins and polypeptides present naturally in the body.

Take over from: D. L. Nelson, M. M. Cox :LEHNINGER. PRINCIPLES OF BIOCHEMISTRY Fifth edition

b-sheet undergoes partial folding, associates partially with the same region in another polypeptide chain (the nucleus of amyloid).

Alzheimer´s disease

Prion protein

1. Globular proteins are compactly folded and coiled.

2. Fibrous proteins are more filamentous or elongated.

3. Peptides Small peptides (containing less than a couple

of dozen amino acids) are called oligopeptides. Long peptides are called polypeptides. Peptides have a "polarity"; each peptide has

only one free a-amino group (on the amino-terminal residue) and one free (non-side chain) carboxyl group (on the carboxy-terminal residue)

Protein Structure

Functional Roles of Proteins

1. Dynamic functiontransportmetabolic controlcontractioncatalysis of chemical transformation

2. Structural functionbone, connective tissue

1. Enzymes (lactate dehydrogenase, DNA

polymerase)

2. Storage proteins (ferritin, cassein, ovalbumin)

3. Transport proteins (hemoglobin, myoglobin,

serum albumin)

4. Contractile proteins (myosin, actin)

5. Hormones (insulin, growth hormone)

6. Protective proteins in blood (antibodies,

complement, fibrinogen)

7. Structural proteins (collagen, elastin,

glycoproteins)

Classification of Proteins by Bioloical Function

Globular proteins Spheroid shapeVariable molecular weightRelatively high water solubilityVariety function roles – catalysts, transporters, control proteins (for the regulation of metabolic pathways and gene expression)

Fibrous proteins Rodlike shapeLow solubility in the waterStructural role in the organism

LipoproteinsComplex of lipids with protein

Glycoproteins Contain covalently bound carbohydrate

Types of Proteins

Globular Proteins

• Globular proteins, such as most enzymes, usually consist of a combination of the two secondary structures.

• For example, hemoglobin is almost entirely alpha-helical, and antibodies are composed almost entirely of beta structures.

Keratin

Fibrilar Proteins

Collagen

Lipoproteins are multicomponent complexes of protein

and lipids. The lipids or their derivatives may be covalently or non-

covalently bound to the proteins. Many enzymes, transporters, structural proteins,

antigens, adhesins and toxins are lipoproteins. Lipoproteins have wide variety function in blood

(transport of lipids from tissue to tissue) and lipid metabolism.

The function of lipoprotein particles is to transport lipids (fats) and cholesterol around the body in the aqueous blood, in which they would normally dissolve

Lipoproteins

GlycoproteinsGlycoproteins have covalently attached sugar molecules at

one or multiple points along the polypeptide chain

Glycoproteins are:• hormones• extracellular matrix proteins• proteins involved in blood coagulation• antibodies• mucus secretion from epithelial cells• protein localized on surface of cells• receptors (transmit signals of hormones or growth

factors from outside environment into the cell)Sugar molecules are:glucose, galactose, mannose, fucose, xylose, N-

acetylglucosamine, N-acetylgalactosamine

Structure-Function Relationship of Protein Families

Human hemoglobin occurs in several forms.

Consist of four polypeptide chains of two different primary structure.

Bind oxygen in the lung and transport the oxygen in blood to the tissues and cells.

Hemoglobin and Myoglobin

Myoglobin is a single polypeptide chain with one oxygen binding site. Binds and release oxygen in

cytoplasm of muscle cells.

Hemoglobin and myoglobin molecules each contain a heme prosthetic group. Protein without prosthetic

group is designated as apoprotein. Complete protein is a

holoprotein

Oxygen binding to Fe2+ of heme in hemoglobin

O2 binding cause conformational changes which pulls Fe2+ back

Proximal histidine binding pulls Fe2+ above the plane of porphyrine ring

Myosin – thick filament of the muscle Actin – thin filament of the muscle G-actin (globular actin) F-

actin (fibrilar actin) Tropomyosin TroponinOne of the biologically important properties of myosin is its ability to

combine with actin to generate muscle contraction.

Contractile Elements of Muscles

Integral membrane proteins

Peripheral membrane proteins

Channels and poresErythrocyte membrane

Diagram of a voltage-sensitive sodium channel α-subunit. G - glycosylation, P- phosphorylation, S - ion selectivity, I - inactivation, positive (+) charges in S4 are important for transmembrane voltage sensing.

Biological Membrane Proteins

Proposed model for insertion of the b2

adrenergic receptor in the cell membrane

1. b-polypeptide stretch extendings from a-helix.

2. Seven membrane-spanning domains.

3. Recognize catecholamines, principally norepinephrine.

Hormone activates receptor.Hormone-receptor mediated

stimulation of intracellular signalling cascade.

Membrane Receptors

Regulatory proteins binding to DNA sequence can promote either an activation or repression of the rate of gene transcription into mRNA

Helix-turn-helix binding proteins The zinc finger motif The leucine zipper motif

The zinc finger motif

Helix-turn-helix motif

DNA Binding Proteins