Structure of proteins

Transcription

1 Structure of proteins Presented by Dr. Mohammad Saadeh The requirements for the Pharmaceutical Biochemistry I Philadelphia University Faculty of pharmacy

2 Structure of proteins The 20 a.a commonly found in proteins are joined together by peptide bonds. Increasing complexity of protein structure is best analyzed by considering four organizational levels, namely, primary, secondary, tertiary, and quaternary. (Figure 2.1).

3 I. Primary structure of proteins The sequence of amino acids in a protein is called the primary structure of the protein. Genetic diseases result in proteins with abnormal amino acid sequences, which cause improper folding and loss or impairment of normal function. If the primary structures of the normal and the mutated proteins are known, this information may be used to diagnose or study the disease.

4 I. Primary structure of proteins A. Peptide bond In proteins, amino acids are joined covalently by peptide bonds, which are amide linkages between the α- carboxyl group of one amino acid and the α-amino group of another. Peptide bonds are not broken by heating or high concentrations of urea. Prolonged exposure to a strong acid or base at elevated temperatures is required to hydrolyze these bonds nonenzymically.

5 I. Primary structure of proteins A.1. Naming of the peptide The free amino end (N-terminal) of the peptide chain is written to the left and the free carboxyl end (C-terminal) to the right. Therefore, 1. all amino acid sequences are read from the N- to the C-terminal end of the peptide. 2. All amino acid residues have their suffixes (-ine, -an, -ic, or -ate) changed to -yl, with the exception of the C-terminal amino acid. example, a tripeptide composed of an N-terminal valine, a glycine, and a C terminal leucine is called valylglycylleucine.

6 I. Primary structure of proteins A.2. Characteristics of the peptide bond The peptide bond has a 1. Partial double-bond character 2. shorter than a single bond, and is rigid planar. 3. Trans configuration. 4. Uncharged but polar. the configurations of the peptide bond is generally a trans bond in large part because of steric interference of the R- groups when in the cis position.

7 I. Primary structure of proteins A.3. Polarity of the peptide bond The C=O and NH groups of the peptide bond are uncharged, and neither accept nor release protons over the ph range of Usally ionized groups present in the side chains of the constituent amino acids. The C=O and NH groups of the peptide bond are polar, and are involved in hydrogen bonds, for example, in α-helices and β-sheet structures.

8 II. Secondary structure of proteins The regular arrangements of amino acids that are located near to each other in the linear sequence. These arrangements are termed the secondary structure of the polypeptide (not straight). The α helix, β-sheet, and β-bend (β-turn) are examples of secondary structures.

9 II. Secondary structure of proteins A. α-helix α-helix is the most common. - It is a spiral structure, consisting of a tightly packed, coiled polypeptide backbone core. - The side chains extending outward from the central axis to avoid interfering sterically with each other. - example, the keratins (α-helical), is related, to fibrous proteins (hair, skin) and their rigidity is determined by the number of disulfide bonds between the constituent polypeptide chains. myoglobin, is also highly α-helical.

10 II. Secondary structure of proteins A. α-helix 1. Hydrogen bonds An α-helix is stabilized by extensive hydrogen bonding between the peptidebond carbonyl oxygens and amide hydrogens. The hydrogen bonds extend up and are parallel to the spiral from the carbonyl oxygen of one peptide bond to the NH group of a peptide linkage four residues ahead in the polypeptide.

11 II. Secondary structure of proteins A. α-helix 2. Amino acid per turn Each turn of an α-helix contains 3.6 amino acids. Thus, amino acid residues spaced three or four residues apart in the primary sequence.

12 II. Secondary structure of proteins A. α-helix 3. Amino acid that disrupt an α-helix 1. Proline (secondary amino group) is not geometrically compatible with the right-handed spiral of the α-helix. 2. Large numbers of charged amino acids (for example, glutamate, aspartate, histidine, lysine, or arginine) disrupt the helix by forming ionic bonds. 3. Amino acids with bulky side chains, such as tryptophan, valine or isoleucine, that branch at the β-carbon can interfere with formation of the α- helix.

13 II. Secondary structure of proteins B. β-sheet The β-sheet is another form of secondary structure. The surfaces of β- sheets appear pleated, figure A. 1. Comparison of a β-sheet and an α- helix: β-sheets are composed of two or more peptide chains (β-strands). Note, in β-sheets the hydrogen bonds are to the polypeptide backbone (Figure A).

14 II. Secondary structure of proteins B. β-sheet 2. Parallel and antiparallel sheets: polypeptide chains that are arranged either antiparallel to each other (with the N- terminal and C-terminal ends of the β- strands alternating as shown in Figure2.7.B), or parallel to each other (with all the N-termini of the β strands together as shown in Figure C). β-sheet can also be formed by a single polypeptide chain folding back on itself (Figure2.7C). The hydrogen bonds are intrachain bonds of the β-sheet. Globular proteins, β-sheets.

15 II. Secondary structure of proteins C. β-bends (reverse turns, β-turns) β-bends reverse the direction of a polypeptide chain, helping it form a compact, globular shape and found on the surface of protein, and often include charged residues. β-bends are generally composed of four amino acids, one of which may be proline that causes a kink in the polypeptide chain. Glycine, is also found in β-bends. β-bends are stabilized by the formation of hydrogen and ionic bonds.

16 II. Secondary structure of proteins D. Nonrepetitive secondary structure The polypeptide chain is described as having a loop or coil conformation. not random, but rather simply have a less regular structure than those described above. [Note: The term random coil refers to the disordered structure obtained when proteins are denatured

17 II. Secondary structure of proteins E. Supersecondary structures (motifs) Supersecondary structures are produced by packing side chains from adjacent secondary structural elements close to each other. for example, α-helices and β-sheets that are adjacent (Figure 2.8).

18 III. Tertiary structure of globular proteins The primary structure of a polypeptide chain determines its tertiary structure. Tertiary refers both to the folding of domains and the final arrangement of domains in the polypeptide. The structure of globular proteins in aqueous solution is compact, with a high-density (close packing) of the atoms in the core of the molecule.

19 III. Tertiary structure of globular proteins A. Domains Domains are the fundamental functional, threedimensional structural and units of polypeptides. Polypeptide chains that are greater than 200 amino acids in length generally consist of two or more domains. The core of a domain is built from combinations of supersecondary structural elements (motifs). Domain has the characteristics of a small, compact globular protein that is structurally independent of the other domains in the polypeptide chain.

20 III. Tertiary structure of globular proteins B. Interactions stabilizing tertiary structure four types of interactions cooperate in stabilizing the tertiary structures of globular proteins. 1. Disulfide bonds: (Figure 2.9). Disulfide bind between two cysteines contributes to the stability of the threedimensional shape of the protein, and prevents it from becoming denatured in the extracellular environment. For example, many disulfide bonds are found in immunoglobulins that are secreted by cells.

21 III. Tertiary structure of globular proteins B. Interactions stabilizing tertiary structure 2. Hydrophobic interactions: Amino acids with nonpolar side chains tend to be located in the interior of the polypeptide (Figure 2.10). Amino acids with polar or charged side chains tend to be located on the surface in contact with the polar solvent.

22 III. Tertiary structure of globular proteins B. Interactions stabilizing tertiary structure 3. Hydrogen bonds: Alcohol groups of serine and threonine, can form hydrogen bonds with the oxygen of a carboxyl group or carbonyl group of a peptide bond (Figure 2.11). 4. Ionic interactions: Negatively charged groups, such as the carboxylate group (-COO - ) in the side chain of aspartate or glutamate, can interact with positively charged groups, such as the amino group (-NH3 + ) in the side chain of lysine (Figure 2.11).

23 IV. Quaternary structure of proteins The arrangement of single polypeptide (monomeric proteins) subunits is called the quaternary structure of the protein. Subunits are held together by noncovalent interactions, hydrogen bonds, ionic bonds, and hydrophobic interactions (see figure). Subunits function independently of each other, or may work cooperatively, as in hemoglobin.

24 Protein folding Protein folding, which occurs within the cell in seconds to minutes, involve nonrandom, ordered pathways. Peptide folds, secondary structures from driven by the hydrophobic effect these small structures combine to form larger structures. Additional events stabilize secondary structure and initiate formation tertiary structure. Finally, the peptide is fully folded, native (functional) from characterized by a low-energy state (figure 2.12)

25 Denaturation of proteins Protein denaturation results in the unfolding and disorganization the secondary and tertiary structures of proteins without the hydrolysis of peptide bonds (see figure). Denaturing agents include heat, organic solvents, mechanical mixing, strong acids or bases, detergents, and ions of heavy metals such as lead and mercury. Denaturation under ideal conditions, be reversible, therefore the protein refolds when the denaturing agent is removed. Most denaturation of proteins be disordered (irreversible). Denatured proteins are often insoluble and precipitate from solution.

26 Role of chaperones in protein folding proteins, folding is a facilitated process that requires a specialized group of proteins, 1. molecular chaperons, 2. adenosine triphosphate hydrolysis. The chaperons, also known as heat shock Proteins (Hsp) interact with the polypeptide at various stages during the folding process. Some chaperones bind hydrophobic region of an extended polypeptide and are important in keeping the protein unfolded until its synthesis is completed (for example, Hsp 70).

27 Protein misfolding Protein folding is a complex process. The deposits of misfolded proteins are associated with a number of diseases. A. Amyloid disease Misfolding of proteins may occur spontaneously, or by a mutation in a particular gene, which produces an altered protein. Abnormal proteolytic cleavage of normal protein leads to the formation of long, fibrillar protein assemblies consisting of β- pleated sheets. Accumulation of these insoluble, spontaneously aggregating proteins, called amyloids, that cause a parkinson and particularly Alzheimer disease. The amyloid plaque that accumulates in Alzheimer disease is amyloid β (Aβ), an extracellular peptide containing amino acid residues (β-pleated, nonbranching fibrils).

28 Protein misfolding Alzheimer factors 1. The Aβ that is deposited in the brain in Alzheimer disease is derived by enzymic cleavage (secretases) from the larger amyloid precursor protein, a single transmembrane protein expressed on the cell surface in the brain and other tissues (Figure 2.13). Most cases of Alzheimer disease are not genetically based, although at least 5 10% of cases are familial. 2. A biologic factor of Alzheimer disease is the accumulation of neurofibrillary tangles inside neurons. A tangled fibers is an abnormal form (hydrophosphorylated and insoluble) of the tau (τ) protein, which, in its healthy version, helps in the assembly of the microtubular structure. The defective τ appears to block the actions of its normal counterpart.

29 Protein misfolding, Alzheimer disease

30 Protein misfolding B. Prion disease Prion protein=normal protein; prion=abnormal protein (almost in brain). The prion protein (PrP) has been strongly implicated as the causative agent of transmissible spongiform encephalopathies (TSEs), including Creutzfeldt-Jakob disease in humans, scrapie in sheep, and bovine spongiform encephalopathy in cattle (popularly called mad cow disease ). the agent causing scrapie in sheep was associated with a single protein species. This infectious protein is designated PrP Sc (Sc=scrapie). PrP Sc is highly resistant to proteolytic degradation, and tends to form insoluble aggregates of fibrils, similar to the amyloid found in some other diseases of the brain.

31 Protein misfolding B. Prion disease Noninfectious PrP c contain α-helix are replaced by PrP sc β-sheets in the infectious form (Figure 2.14). PrP c + PrP sc PrP sc + PrP sc The infective agent is an altered of a normal protein, which acts as a template for converting the normal protein to the pathogenic conformation. The TSEs are invariably fatal, and no treatment is currently available that can alter this outcome.

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