Denaturation of protein

INTRODUCTION
WHAT IS DENATURATION?
SOME EXAMPLES OF DENATURATION
THERMAL DENATURATION.
HOW DENATURATION OCCURES AT THE LEVEL OF PROTEIN STRUCTURE
NUCLIEC ACID DENATURATION
DENATURANTS
CONSEQUENSES OF DENATURATION
CONCLUSION
REFERENSE

1. DENATURATION OF PROTEIN By KAUSHAL KUMAR SAHU Assistant Professor (Ad Hoc) Department of Biotechnology Govt. Digvijay Autonomous P. G. College Raj-Nandgaon ( C. G. )

2. CONTENTS- INTRODUCTION WHAT IS DENATURATION? SOME EXAMPLES OF DENATURATION THERMAL DENATURATION. HOW DENATURATION OCCURES AT THE LEVEL OF PROTEIN STRUCTURE NUCLIEC ACID DENATURATION DENATURANTS CONSEQUENSES OF DENATURATION CONCLUSION REFERENSE

3. INTRODUCTION Denaturation  is a process in which  proteins or   nucleic acids  lose the  tertiary structure  and  secondary structure  which is present in their native state , by application of some external stress or compound such as a strong  acid or   base , a concentrated  inorganic   salt, an organic solvent (e.g.,  alcohol  or  chloroform ), or  heat . . If proteins in a living cell are denatured, this results in disruption of cell activity and possibly cell death.

4. WHAT IS DENATURATION? Denaturation is a phenomenon that involves transformation of a well-defined, folded structure of a protein, formed under physiological conditions, to an unfolded state under non-physiological conditions. Denatured proteins can exhibit a wide range of characteristics, from loss of solubility to communal aggregation. Communal aggregation is the phenomenon of aggregation of the hydrophobic proteins to come closer and form the bonding between them, so as to reduce the total area exposed to water .

5. Fig:The protein "albumin" in egg white undergoes denaturation and loss of solubility when the egg is cooked. (Bottom) Paperclips provide a visual analogy to help with the conceptualization of the denaturation process .

6. When and How are Proteins Denatured? At Very High or Low pH . At Very High Temperatures . By Heavy Metal Ions . By Small Polar Molecules.

7. Denaturation at HIGH or LOW pH

8. DENATURATION BY REDUSING AGENTS-

9. Denaturation by Small Polar Molecules- Urea ( CO(NH 2 ) 2 ) in concentrated solution will denature proteins . It disrupts the Hydrogen Bonds . This causes complete denaturation

10. THERMAL DENATURATION- Rate of denaturation depends on the temperature As T is increased – Affect interactions of tertiary structure – Increased flexibility _ reversible – H-bonds begin to break _ water interaction – Increased water binding – Increased viscosity of solution – Structures different from native protein Upon cooling – Aggregation – Loss of solubility Water content affects heat denaturation Other consequences – Splitting of disulfide bonds – Chemical alterations of amino residues – Crosslinks

11. SOME EXAMPLES OF DENATURATION- A classic example of denaturing in proteins comes from  egg whites , which are largely egg  albumins  in water . Fresh from the eggs, egg whites are transparent and liquid. Cooking the  thermally unstable  whites turns them opaque, forming an interconnected solid mass . The same transformation can be effected with a denaturing chemical. Pouring egg whites into a beaker of  acetone  will also turn egg whites translucent and solid. The skin that forms on  curdled  milk is another common example of denatured protein .

12. HOW DENATURATION OCCURES AT THE LEVEL OF PROTEIN STRUCTURE- In  quaternary structure   denaturation , protein sub-units are dissociated and/or the spatial arrangement of protein subunits is disrupted. Tertiary structure   denaturation involves the disruption of: Covalent  interactions between amino acid side-chains (such as  disulfide bridges  between  cysteine  groups ) Noncovalent  dipole-dipole interactions between polar amino acid side-chains (and the surrounding solvent ) Van der Waals (induced dipole) interactions between nonpolar amino acid side-chains.

13. In  secondary structure   denaturation , proteins lose all regular repeating patterns such as alpha-helices and beta-pleated sheets, and adopt a random coil configuration. Primary structure , such as the sequence of amino acids held together by covalent peptide bonds, is not disrupted by denaturation . CONTINUE......

14. Reversibility and irreversibility- In many case (unlike egg whites), denaturation is reversible (the proteins can regain their native state when the denaturing influence is removed ). This process can be called renaturation .  This understanding has led to the notion that all the information needed for proteins to assume their native state was encoded in the primary structure of the protein, and hence in the  DNA  that codes for the protein, the so called " Anfinsen's   thermodynamic hypothesis”

15. DENATURANTS- Acidic protein denaturants include: Acetic acid Trichloroacetic acid 12% in water Sulfosalicylic acid Solvents Most organic solvents are denaturing, including: Ethanol Methanol

16. Cross-linking reagents Cross-linking agents for proteins include: Formaldehyde Glutaraldehyde Chaotropic agents Chaotropic agents include: Urea 6 - 8 mol/l Guanidinium chloride 6 mol/l Lithium perchlorate  4.5 mol/l CONTINUE……

17. Disulfide bond reducers- Agents that break disulfide bonds by reduction include: 2-Mercaptoethanol Dithiothreitol TCEP ( tris (2-carboxyethyl) phosphine ) CONTINUE……..

18. Loss of enzymatic activity (death) Destruction of toxins Improved digestibility Loss of solubility Changes in texture CONSEQUENSES OF DENATURATION-

19. CONCLUSION- The conversion of a biologically functional molecule into a non-functional form There are many denatured states but one native state Proteins can regenerate to their native state but slowly. Most biological substrates lose their biological function when denatured. For example,  enzymes  lose their  activity , because the substrates can no longer bind to the  active site , and because amino acid residues involved in stabilizing substrates'  transition states  are no longer positioned to be able to do so