Biomolecules (Proteins)

1. Biomolecules(Proteins) L.V. Athiththan Dept. of Biochemistry

2. Objectives • Sources & function of protein • Chemical & 3D structure of biological molecule • Forces stabilizing protein molecule • Denaturation of proteins & factors enhancing it • Changes in structure can lead to various diseases • Qualitative & quantitative estimation of amino acid & proteins • pI

3. Introduction • When there is something to do, it is a protein that does it.

4. Role of Proteins • It makes up the muscles, tendons, organs, glands, enzymes, hormones, nails, and hair. • It is needed for growth, repair and maintenance of all cells • Next to water, protein makes up the greatest portion of our body weight

5. What is protein? • Basically proteins are made up of amino acids (several amino acid forming long chain) It is found in every cell mainly in muscle tissues of our body • It is present in many of the foods that we eat.

6. Amino acid • These are the chemical units or "building blocks" of the body that make up proteins. • Amino Acids that must be obtained from the diet are called "Essential Amino Acids" other Amino Acids that the body can manufacture from other sources are called "NonEssential Amino Acids."

7. Essential amino acids A.A that cannot be synthesized in our body at required amount is known as essential a.a e.g. Histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine , tryptophan, valine. Arginine mainly in children only?

8. STRUCTURE OF AMINO ACID

9. STRUCTURE OF AMINO ACID • Each a.a has • a carboxyl group • a primary amino group (proline 2ry ) • & a distinctive side chain “R group” • At physiological pH the carboxyl group is negatively charged (COO-)& the amino group (NH3+) is protonated(Zwitter ion)

10. Optical properties of amino acid • The α-carbon of a.a is a chiral carbon (attached to four different groups) • It has a optically active carbon atom • Glycine is exception • They have the mirror image form - D & L isomerism

12. The two forms in each pair are termed (sterioisomers/optical isomers/ enantiomers) • All amino acid found in protein are of L-configuration

13. Classification of amino acid (I) A.a acid with nonpolar side chain e.g. glycine, alanine, valine, phenyl alaninemethionine, leucine, etc. Proline contains an imino group

14. Classification of amino acid (II) A.a with uncharged polar side chains Serine, threonine, tyrosine, asparagine, cysteine, glutamine

15. Classification of amino acid (III) Amino acid with acidic side chain Aspartic acid, glutamic acid (IV) Amino acid with basic side chains Histidine, lysine, arginine

17. Abbreviation & symbols of a.a • Each a.a has a 3 letter abbreviation and one-letter codes e.g. Cysteine = Cys = C (Unique first letter) Histidine = His = H Isoleucine = Ile = I

18. If more than one a.a begins with a particular letter ? • Most common a.a has a priority • Alanine = Ala = A • Similar sounding names • Arginine = Arg = R • Letter close to initial

19. Isoelectric point (pI) • The pI is sometimes abbreviated to IEP, is the pH at which a particular molecule or surface carries no net electrical charge (or the negative and positive charges are equal) • Amphoteric molecules (zwitterions) contain both positive and negative charges depending on the functional groups present in the molecule.

20. Isoelectric point (pI) • The net charge is affected by pH of the environment & can become more positively or negatively charged due to the loss or gain of protons (H+). • pI value can affect the solubility of a molecule i.e. At pI the molecule has the minimum solubility

21. Isoelectric point (pI) • At a pH below their pI, proteins carry a net positive charge; above their pI they carry a net negative charge. • Proteins can thus be separated according to their isoelectric point using a technique called isoelectric focusing.

22. Calculating pI values • For an a.a with only one NH2and one COOH group, the pI can be calculated from the pKa’s of this molecule.

23. Calculating pI values • For amino acids with more than two ionizable groups, such as lysine, the same formula is used • But the two pKa's used are those of the two groups that lose and gain a charge from the neutral form of the amino acid.

25. Peptide bond • Two a.a can be joined covalently in a condensation reaction. • These are amide linkages between the α-carboxyl group of one a.a and the α-amino group of the other • This reaction is catalyzed by the ribosome in a process known as translation.

28. Peptide bond • Peptide bonds are not broken under normal denaturing conditions. • Peptide bond • has a partial double bond character (Shorter than a single bond ) • Is rigid and planar • Exist in trans configuration • Is uncharged but polar • Can be freely rotated

29. Peptide bond • Peptide bonds are uncharged and neither accepts nor release protons Polarity depends on the N-terminal amino group, C-termibnal carboxy group and any ionized chains present in the side chains

30. Peptide bond • The peptide bond is in fact planar due to the delocalization of the electrons from the double bond. • The rigid peptide bond between C1 and N is always closer to 180 degrees.

31. Naming the peptide bond • Free N terminal is written to the left • Free Carboxyl terminal to the right • A.a are read from the N to C terminal • A.a residue having the suffix ine, an, ic, ate are changed to “yl” except the c-terminal

33. Seryltyrosylcysteine

34. Structure Organization Of Proteins • Mainly four organizational level • Primary, Secondary, Tertiary, Quaternary. • The complexity increases • Ranges from simple proteins to multifunctional proteins

36. Primary structure (Iry) • Sequence of a.a in a protein (joined by peptide bonds) Includes the location of disulfide bond • Deviation and abnormality in the a.a sequence may result in improper folding and loss of normal function

38. Secondary structure (IIry) • The conformation of the polypeptide backbones of proteins Commonly they form a regular arrangements with a.a close to each other • The arrangements are due to the partial double bond character of the peptide bond

39. Secondary structure (IIry) • Secondary structure includes α-helix, β-sheet and β-bend • Hydrogen bonds and Vander Waals force stabilize the secondary structure mainly the α-helix

40. Alpha helix (α-helix) • A common motif in the IIry structure of proteins • The α-helix is a right- or left-handed coiled conformation • Resembles a spring, in which every backbone N-H group donates a H bond to the backboneC=O group of the a.a four residues earlier ( hydrogen bonding).

42. Alpha helix (α-helix) • Spiral structure with tightly packed coiled polypeptides, where the side chain is protruded out • It has the lowest energy and is the most stable form (stability increases with hydrogen bond formation)

43. Alpha helix (α-helix) • Each turn contains 3.6 a,a • Some a.a disturb the helix • proline may insert a kink, charge a.a form ionic bond and disturb the arrangement, and bulk side chain a.a may also interfere

44. Beta sheet (β-sheet) • Unlike the alpha helix β-chain are composed of two or more peptide chains (strands) or segments of peptide chain • All the peptide bond component are involved in hydrogen bonding • The hydrogen bond is perpendicular

45. Beta sheet (β-sheet) • Hydrogen bonds may be inter or intra chain bonds • They may be parallel (C–terminals on the same side or antiparallel (C-terminals on the opposite side)

46. ANTIPARALLEL PARALLEL

47. C terminal C terminal N terminals

48. β - bends/Reverse turns • Predominant protein type in globular proteins • Compact structure • Predominantly found in cytosol & surface of protein molecule • Connect strands of anti-parallel β-sheets • Contain mainly proline (kink), glycine(smallest a.a)

49. β-bend

50. Other secondary structures • Non repetitive secondary structure • Some polypeptides may have a loop or coil and have less regular structure (“random coil”) • Super secondary structure • Combines all IIry structure & assembles together • Side chains of adjacent IIry structure elements assemble close to each other & folds into a protein