PROTEINSProteins are probably the most important class ofbiochemical molecules, although of course lipidsand carbohydrates are also essential for life.Proteins are the basis for the major structuralcomponents of animal and human tissue.
Classification of Some Proteins and their FunctionsClass of Protein Function in the body ExamplesStructural Provide structural Collagen components KeratinContractile Move muscles Myosin ActinTransport Carry essential Hemoglobin substances throughout Lipoprotein the bodyStorage Store nutrients Casein FerritinHormone Regulate body Insulin metabolism and nervous Growth hormone systemEnzyme Catalyze biochemical Sucrase reactions in the cells TrypsinProtection Recognize and destroy immunoglobulins foreign substances
Question• Classify each of the following proteins according to its function____________1. hemoglobin(oxygen carrier in the blood)____________2. collagen(a major component of tendons and cartilage____________3. keratin( a protein found in hair)____________4. amylase( an enzyme that hydrolyze starch____________5. casein(milk protein)
AMINO ACIDS Overview• Amino Acids are the chemical units or "building blocks" of the body that make up proteins. Protein substances make up the muscles, tendons, organs, glands, nails, and hair. Growth, repair and maintenance of all cells are dependent upon them. Next to water, protein makes up the greatest portion of our body weight.
Amino acids are the basic structural units of proteins.An alpha-amino acid consists of anamino group, a carboxyl group, ahydrogen atom, and a distinctive Rgroup bonded to a carbon atom, whichis called the alpha-carbon because it isadjacent to the carboxyl (acidic) group.An R group is referred to as a sidechain.(
• Contain 2 functional groups:amino (-NH2) groupcarboxylic acid (-COOH) group.R group = amino acid side chain. This is whatdistinguishes one amino acid from another.• If the amino group, carboxylic group, R-group, and hydrogen are bonded to a central carbon atom (α-carbon), they are termed α-amino acids.• Only 20 different amino acids are present in the human Their names are abbreviated with 3 letters.• All of the α-amino acids (except glycine) are chiral because the a-carbon is attached to 4 different groups.
Chiral Carbon• A carbon atom that is bonded to four different atoms or groups of atoms.• The α carbon is chiral or asymmetric ( 4 different groups are attached to the carbon; exception is glycine.)
CLASSIFICATION OF AMINO ACIDSI. BASED ON THE NATURE OF R A. non-polar or hydrophobic R 1. R is an aliphatic radical e.g. ala, val, ile, leu 2. R is aromatic e.g. phe, trp B. polar R but uncharged 1. R is aromatic e.g. tyr, trp, phe 2. R has an –OH group e.g. ser, thr 3. R has an –S-group, e.g. cys, met 4. R has an amide group, e.g. asn, gln 5. R is an H, e.g. gly
C. R is positively charged, e.g. lys, arg, his D. R is negatively charged, e.g. asp, glu E. an imino acids, e.g. proII. ACCORDING TO ACID-BASE PROPERTIES A. neutral amino acids, e.g. ala, val B. acidic amino acids, e.g. glu, asp C. basic amino acids, e.g. arg, lysNOTE: acidic amino acids are negatively charged because theyhave two carboxylic groups(-COOH) and one amino group (-NH2). Basic amino acids are positively charged because theyhave two amino groups (-NH2 ) and only one carboxylic radical (-COOH). Neutral amino acids have one amino group and onecarboxylic group.
PROLINE• Proline shares many properties with the aliphatic group.• Proline is formally NOT an amino acid, but an imino acid(is any molecule that contains both imino (>C=NH) and carboxyl (-C(=O)-OH) functional groups)• Nonetheless, it is called an amino acid. The primary amine on the α carbon of glutamate semialdehyde forms a Schiff base with the aldehyde which is then reduced, yielding proline.
• In addition to amino acids found in proteins, some amino acids are known not to occur in proteins. Examples of non-protein amino acids are: beta alanine (a building block of the vitamin, pantothenic acid); ornithine, an intermediate of the urea cycle; and gamma-aminobutyric acid (GABA), a chemical agent for the transmission of nerve impulses.
α - Amino Acids• All amino acids isolated from proteins, with the exception of proline, have this general structure. Lost proton- carboxylate group Gained proton- protonated Amino group
Carboxylate group• a carboxyl group that has lost a proton; conjugate base form (-COO-)Protonated amino group• An amino group that has gained a proton; conjugate acid form (NH+3)
zwitterion• The dipolar form of an amino acids and has a net charge of zero. Two isomeric forms of an amino acid. The isomer on the right is a zwitterion. Amino acid Zwitterion (dipolar ion)
ZWITTERION• Amino acid physical properties indicate a "salt-like" behavior. Amino acids are crystalline solids with relatively high melting points, and most are quite soluble in water and insoluble in non-polar solvents. In solution, the amino acid molecule appears to have a charge which changes with pH.
ZWITTERION• An intramolecular neutralization reaction leads to a salt- like ion called a zwitterion. The accepted practice is to show the amino acids in the zwitterion form.• (1) The carboxyl group can lose a hydrogen ion to become negatively charged.• (2) The amine group can accept a hydrogen ion to become positively charged.
ZWITTERION• At pH of 7, amino group is protonated (-NH3+) and carboxyl group is ionized (COO-). The amino acid is called a zwitterion.• At the pH of most bodily fluids, the carboxyl group will loose H+ and the amino groups accepts an H+• This is a dipolar (DIPOLE ION)form of the amino acid
ISOELECTRIC POINT• At a certain pH called the isoelectric point (pI), the positive and negative charges on the amino acid are equal and the overall charge is zero. • In a solution that is more basic than pI, the -NH3+ group will loose H+, and the overall charge will be negative. • In a solution that is more acidic than pI, the -COO- accepts an H+, and the overall charge will be positive.
Acid - Base Properties of Amino Acids:• 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.
• 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.
AMINO ACID AS ACIDZwitterions for acidic amino acids typically exist at pH values ofabout 3 because the carboxyl group in their side chain mustpick up H+AMINO ACID AS BASEZwitterions for basic amino acids typically exit at pH 7.6-10.8POLAR AND NONPOLAR AMINO ACIDSZwitterions for polar and nonpolar amino acids typically exit atpH values of 5.0-6.0
NEUTRAL SIDE CHAINS• Since an amino acid has 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 then the whole amino acid is neutral.• 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.•
PRINCIPLES OF POLARITY• The greater the electronegativity difference between atoms in a bond, the more polar the bond. Partial negative charges are found on the most electronegative atoms, the others are partially positiveNon-Polar Side Chains:• Side chains which have pure hydrocarbon alkyl groups (alkane branches) or aromatic (benzene rings) are non-polar.• 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 non-polar than alanine; leucine is more non-polar than valine
NONPOLAR SIDE CHAINS• The non-polar amino acids include: alanine, cysteine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, tyrosine and valine.• Non polar side chains consist mainly of hydrocarbon. Any functional groups they contain are uncharged at physiological pH and are incapable of participating in hydrogen bonding
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.
• 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.
Structures of Amino Acids R = any number carbons in a hydrocarbon chain *CHIME plug-in required to view these images. A A Amino b b Acid r r Structure Comments Name e e of R group (red) v. v. Neutral Alanine ala A Non-polar Basic Arginine arg R Polar NeutralAsparagine asn N Polar Aspartic Acidic asp D Acid Polar NeutralCysteine cys C Slightly PolarGlutamic Acidic glu E Acid Polar NeutralGlutamine gln Q Polar
Neutral Glycine gly G Non-polar BasicHistidine his H Polar NeutralIsoleucine ile I Non-polar NeutralLeucine leu L Non-polar BasicLysine lys K Polar NeutralMethionine met M Non-polar Phenyl- Neutral phe F alanine Non-polar NeutralProline pro P Non-polar NeutralSerine ser S Polar
NeutralThreonine thr T Polar Neutral Trypto- trp W Slightly phan polar NeutralTyrosine tyr Y Polar NeutralValine Val V Non-polar
ISOELECTRIC POINT• Solid amino acids have a very high melting points because the zwitterion has the properties of a salt. The ionic charges of the amino acids make them more soluble in water, but not in organic solvents
PROPERTIES OF AMINO ACIDS1. Protein are soluble in water, insoluble in organic solventslike benzene and ether. They have high meting points(above 200 ◦Celsius) and low vapor pressure. They havelarge dipole moments and high dielectric constants. Theseproperties indicate the amino acids are polar. Polar formsof an amino acid are called zwitterions.
PROTEIN SOLUBILITYProtein solubility depends on several factors.1. It is observed that at low concentration of the salt, solubility of the proteins usually increases slightly. This is termed Salting in. But at high concentrations of salt, the solubility of the proteins drops sharply. This is termed Salting out and the proteins precipitate out.2. Precipitation by changing the pH of the protein solution. This effect is due to the different functional groups on a protein. There will be some pH, known as the isoeletric point where the net charge on the protein is zero. This is different for different proteins.3. You can also precipitate proteins by the addition of a non-ionic polymer or metal ions. (ionic strength)
PROTEIN PRECIPITATION• Precipitation is widely used for product recovery of biomolecules especially proteins. The most common type of precipitation for proteins is salt induced precipitation. Precipitation is usually induced by1. addition of a salt or an organic solvent2. by changing the pH to alter the nature of the solution.
• Why Is Protein Precipitation Special?• Proteins are different than most other molecules. The physical structure is just as important as the chemical structure. If the structure of the protein changes, the protein does not have the activity you want. This could even be harmful
PROTEIN DENATURATION• Denaturation of a protein occurs when there is a disruption in any of the bonds that stabilize the secondary, tertiary, or quaternary structures. The primary structure is not affected. Proteins can be denatured by:• · Heat = disrupts hydrogen bonds and hydrophobic attractions between R groups• · Acids and bases = disrupt hydrogen bonds and salt bridges• · Organic compounds = disrupt hydrophobic interactions between R groups• · Heavy metal ions = disrupt disulfide bonds• · Agitation = disrupts hydrogen bonds and hydrophobic interactions between R groups by stretching the polypeptide chain.•
Proteins Can Be DenaturedBoth temperature and pH can change polypeptide shape.a. Examples: heating egg white causes albumin to congeal;adding acid to milk causes curdling. When such proteinslose their normal configuration, the protein is denatured.b. Once a protein loses its normal shape, it cannot performits usual function.The sequence of amino acids, therefore, forecasts theproteins final shape.
Color Reactions of Proteins• A color test for a protein tests for a particular functional group or structural component present in a protein or in a particular amino acid found in proteins.(a) Biuret reaction.(b) Xanthoproteic reactionThe color is produced as the result of the formation of nitro-derivatives of the compounds which contain a benzene ring, forexample, tyrosine.(c) Million’s reaction.The reaction serves as a test for the presence of tyrosine.(d) Sulphur reactionThe precipitate, which is lead sulphide, is formed as the result ofthe decomposition of the cysteine by the alkali.(e) Hopskins Cole ReactionThe color produced is due to the formation of a compound fromthe glyoxylic acid in the reagent and the tryptophan in the protein
Name of the Test Reagent Used Positive results RemarksBiuret Test NaOH, dilute violet + results with CuSO4 polypeptides and proteinsXanthroproteic Conc. H2SO4 AA with benzene Proteins with trp, tyr,Test ring (yellow) pheMillion’s Test Hg(NO3) and Tyrosine (red) + phenolic compounts Hg(NO2)2Sulphur Test Lead acetate, Gray or black + lead sulphide, dissolved with precipitate formed as the result of NaOH the decomposition of the cysteine by the alkali.Hopkins Cole Test Glyoxylic acid, Tryptophan (violet + with any compound sulfuric acid ring) with indole ringNinhydrin Test ninhydrin Free-NH2 group + results given by NH3, (blue) primary amines, amino acids, peptides, and proteins
Indole-an aromatic heterocyclic organic compound.It has a bicyclic structure, consisting of a six-membered benzene ringfused to a five-membered nitrogen-containing pyrrole ring. Indole is apopular component of fragrances and the precursor to manypharmaceuticals. Compounds that contain an indole ring are calledindoles. The indolic amino acid tryptophan is the precursor of theneurotransmitter serotonin.
PROPERTIES OF AMINO ACIDS2. Except for glycine, the alpha carbon is asymmetric orchiral and therefore capable of optical isomerism. Thenumber of possible isomers is given by the formula 2nwhere n is the number of chiral carbons3. Aromatic amino acids absorb light in the ultravioletregion of the EMR. This property is used for estimatingprotein content. E.g. phe absorbs at 260 nm, trp at 280 nm,tyr at 275 nm4. Naturally occurring amino acids have an L-configuration.By convention, L-amino acids are written with NH3 to theleft of the alpha carbon and the –COO on top.5. Amino acids may be acidic, basic, or neutral.
Stereoisomers Stereochemistry is the study of the 3-dimensional structure of molecules. Isomers are molecules with the same chemical formula and often with the same kinds of bonds between atoms, but in which the spatial arrangement of atoms differs. Isomers are grouped into two broad classes. (Most of the non- substituted cycloalkanes have conformational isomers, or diastereomers also known as conformers.)
If you orient the molecule so that you look along itfrom the COO− group to the NH3+ group, themethyl (R) group can extend out to the left,forming L (shown below on the left) or to the -alanineright, forming D (on the right). -alanine
• L amino acids are used exclusively for protein synthesis by all life on our planet.• Does it really matter? Yes.• The function of a protein is determined by its shape.• A protein with a D amino acid instead of L will have its R group sticking out in the wrong direction.• Many other kinds of organic molecules exist as enantiomers. Usually only one form is active in biological systems. For example, if one form binds to a receptor protein on the surface of a cell, the other probably cannot.
QUESTIONSI. Write the zwitterion of each of the following amino acids1. Glycine2. Cysteine3. Serine4. Alanine5. PhenylalanineII. Write the Fischer projection for1. L-serine2. L-cysteine
III. List all amino acids with non-polar side chains.IV. Rank the following according to increasing non-polarityi.e. 1 = least non-polar, 4 = most non-polar.A. leuB. pheC. valD. alaV. List all amino acids by abbreviation which are consideredsomewhat polar.
VI. Rank the following amino acids by increasing polarity.i.e.1 = more non-polar.A. serB. gluC. aspD. lysE. alaF. glnVII. Which amino acid is most insoluble in water: isoleucineor alanine ? Explain.VIII. Which amino acid is most soluble in water: lys orser?Explain.
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