Understanding Protein Structure: Primary, Secondary, Tertiary, and Quaternary Levels, Transcriptions of Biochemistry

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[BIOCHEM LEC] Unit 2 Proteins: Peptides PEPTIDES AND ITS STRUCTURES Levels of protein organization © Primary (1°) o Simplest level of protein structure o Amino acid sequence in protein structure and function o The amino acid composition o Thesequence of amino acids from N terminal to the C terminal o Stabilized by peptide bonds o Amino acid sequence is a form of genetic information o Unique amino acid sequence makes the protein distinctive from others = It defines the structures, shape and function of protein = The sequence is dictated by the DNA base sequence ina gene * Errors in the DNA may result to erroneous, non-functional protein o Each chain has its own set of amino acids, assembled in a particular order o Wrong sequence of amino acids can create a different fold and structure leading to the inability to function well o The protein will not function well if it will not reach its “native” form o Ex: sickle cell anemia = The glutamic acid that is normally the sixth amino acid of the hemoglobin B chain (one of two types of protein chains that make up hemoglobin) is replaced by a valine = The difference between a normal hemoglobin molecule and a sickle cell molecule is just 2 amino acids out of the approximately 600 * glutamic acid-to-valine amino acid change makes the hemoglobin molecules assemble into long fibers (crescent shapes) HHO Io threonine — proline —N—C — O— Ilys Normal aller acs Noma ino acid HH o swosttten threonine — proline — N—C — 0 — lys s ES OTSO valine Sickle cell hemoglobin Gene Genes DNA molecule Gene2 DNAstrand [nme RNA —S— Ss Coron rranstanbif | | | Polypeptide Aminoacid Secondary (2°) o Refers to local folded structures that form within a polypeptide due to interactions between atoms of the backbone (specifically R groups are not included in the interaction) o H-bonding formed within atoms of peptide bond o Stabilized by hydrogen bonds between C=O of one peptide bond (residue) and the N-H of another o Primary structure dictates the secondary structure o Amino acids such as tryptophan, tyrosine, and phenylalanine, which have large ring structures in their R groups, are often found in B pleated sheets o Proline: “helix breaker"; its unusual R group (which bonds to the amino group to form a ring) creates a bend in the chain and is not compatible with helix formation; less likely to be found in a-helix and pleated sheets ARTICONA — 2FMT [BIOCHEM LEC] Unit 2 Proteins: Peptides o Types of secondary structures: " a-Helix > the oxygen atom of the carbonyl! (C=O) of one amino acid is hydrogen bonded to the H of the amino group (IN-H) of an amino acid that is four down the chain. Formation of H-bond of n and n+4 amino acid in a-helix > E.g. the carbonyl of amino acid 1 would forma hydrogen bond to the N- Creates an H-bond between O- of one peptide bond H of amino acid 5 to the H atom of the 4" peptide bond > N-H groups: point in the same direction, o Helix Breakers almost parallel to the axis of the helix > C=O groups: point in the opposite = Reason for some polypeptides to form a- direction alternately, almost parallel to nelces the avis of the helix * Proline: the rotation around the N-aC > groups: pointed outward of the helix bond is restricted because it is part of the + B-Pleated sheets ring; N has no H to participate in H bonds > carbonyl and amino groups from = Glycine: has more conformation flexibility adjacent chains point toward each due to its R-group; it supports other other and are inthe same plane conformations (6.9. coll or bend) > forms Hebonding o Electrostatic repulsion/attraction > Rgroups extend above and below the = Between successive amino acid residues plane of the sheet (has the same charge consecutively) > Parallel: pointing in the same direction © Bulkiness (steric strain) (meaning that their N- and C-termini * Adjacent R-groups match up) o Random coils > Antiparallel: pointing in opposite : Irovlara’ uniave conformation directions (meaning that the N-terminus of one strand is positioned next to the PARALLEL B-SHEET C-terminus of the other) NJ = Bbends or reverse bends or B-turns or tight E turns rr ty > Proline (provides less flexibility, hence No starting the turn) and glycine (greater - flexibility, hence facilitating the turn) a sasihts gh play common roles in turns i *¥ = random coil b Te apa acmerean <° Bestrands e a Parallel sheet run in = other helical structures in ineartebonds (stronger) posit ese ieee had a * Tertiary (3°) o Three-dimensional structure of a polypeptide 0 Covalent and non-covalent interactions within R-groups to form 3D shapes o Achieved due to covalent and non-covalent interactions between or among R-groups o H-bonding, Hydrophobic, lonic bonding = Formation of these bonds depend on the R-groups o Dipole-dipole interaction: * Intermolecular attractions between two molecules = occur when the partial charges formed within one molecule are attracted to an ARTICONA — 2FMT [BIOCHEM LEC] Unit 2 Proteins: Peptides e¢ = Andon the forces of interaction o The forces make the protein fold e The number of folding patterns are large but infinite e Proteins assume general shapes: globular and fibrous Denaturation e ~The process of destroying the native conformation of a protein by chemical or physical means e¢ Some are reversible, while others are permanently damaged Denaturing Agents e Heat: can disrupt hydrogen bonding; in globular proteins, it can cause unfolding of polypeptide chains with the result that coagulation and precipitation may take place ~Z —_;) is, q => RY > ere => Ge ‘acne ante: on e 6M aqueous urea: Disrupts hydrogen bonding @ urea © water e Surface active agents: Detergents such as sodium dodecylbenzenesulfate (SDS) disrupt hydrogen BEFORE SDS scluarged R-groups hnychophobie areas * Reducing agents: 2-Mercaptoethanol (HOCH 2.CH 2 SH) cleaves disulfide bonds by reducing S-S groups to -SH groups. H 0 4» 0 1 i ww cH tow I Hc HC 2HSCH,CH,OH Se - CH,CH,OH e Heavy metal ions: Transition metal ions such as Pb**, Hg#tand Ca form water insoluble salts with -SH groups; Hg 2+ for example forms SHgS ° i ah 1gN. Cw Oe fin NH Bed ipvecpitatey —° e Alcohols: 70% ethanol penetrates bacteria and kills them by coagulating their proteins. It is used to sterilize skin before injections. Tertiary Structure - Hydrogen Bonding ARTICONA — 2FMT [BIOCHEM LEC] Unit 2 Proteins: Peptides Additional Information: Amino Acid __ahelix Reverse turn__B sheet Ala 1.41 0.82 0.72 Arg 1.21 0.90 0.84 Asn 0.76 1.34 0.48 Asp 0.99 1.24 0.39 Cys 0.66 0.54 1.40 Gin 1.27 0.84 0.98 Glu 1.59 1.01 0.52 Gly 0.43 177 0.58 His 1.05 0.81 0.80 le 1.09 0.47 1.67 Leu 1.34 0.57 1.22 Lys 1.23 1.07 0.69 Met 1.30 0.52 144 Phe 1.16 0.59 1.33 Pro 0.34 1.32 0.31 Ser 0.57 1.22 0.96 Thr 0.76 0.96 147 Tip 1.02 0.65 1.35 Tyr 0.74 0.76 1.45 Val 0.90 0.41 1.87 Relative tendencies of each amino acid to be ina secondary structure. Higher values indicate greater tendency Amino Acid Hydropathy Scores Amino Acid One Letter Code Hydropathy Score Isoleucine | 45 Valine v 42 Leucine L 38 Phenylalanine F 28 Cysteine c 25 Methionine = M 19 Alanine A 18 Glycine G 04 Threonine = T 07 Tryptophan W 09 Serine s 08 Tyrosine Y 13 Proline Pp 1.6 Histidine H 32 Glutamic acid E 35 Glutamine Q 35 Aspartic acid D 35 Asparagine NN 35 lysine kK 39 Arginine R 45 Hydrophobicity of amino acids: lsoleucine being the least polar, arginine being the most polar ACID-BASE PROPERTIES OF PEPTIDES Peptide bond e Amide bond between the a-carboxyl group of one amino acid and the a-amino group of another amino acid o Amino acid residues: what remains on each amino acid when bonded with another o Dipeptide: one peptide, 2 amino acid residues o Tripeptide: 2 peptide bonds, 3 amino acid residues, etc. t if : Hs—CH—C—OH + H—N—CH—Coo" ~ | 1 | H3N—CH—C=N—CH—COO™ e Has considerable C-n double bond character e = Their structure is stabilized by resonance o Rotation of peptide is restricted o Peptide structure is planar tend 3 »-4 a SAAN tie Se VS Sea Samide pune ‘Peptide group © kesonance srvcurest © Toe planer peptide groop. the peptide group. e = Rotations are permitted at the bonds b/w amino group and carbonyl group with the a-carbon o Allows 2 possible conformation: trans (highly favored) and eis conformations uf He wt Se ee \e~ ARTICONA — 2FMT [BIOCHEM LEC] Unit 2 Proteins: Peptides e = N-terminal: has a free amino group at the start/end of the amino acid sequence e C-terminal: has a free carboxyl group Examples: Peptide: DK (D: Asp, K: Lys): Aspattyl lysine Asp pKa: a-NH3+= 9.82, RH=3.86 (c-COOH is not needed because it is connected to the amino group of the other amino acid residue to create a peptide bond) Lys pKa: a-COOH= 2.18, &-NH3+= 10.53 (a-NH3+ is not needed because it is connected to the COOH of the other AA resisue) \\ esd . Cwm h oe ha TEN 2 Ib PRE 4.2 Oa Oo Cow ix-3 db e pHO-] (net charge: +2) o 2(+) charge: 2 NH3+ groups from diff. AA residue o 2 (0) charge: 2 COOH * Add OH- until pH=2.18 (net charge +1) o 50% of COOH of Lys donates a proton becoming COO- o 2(+) charge: 2 NH3+ o 1(0) charge: COOH 1(-) charge: COO- (pKa= 2.18 in Lys) . add OH- until DH 3.86 (net charge: 0) o 50% of B-COOH of Asp donates a proton and becomes COO- o 2(+) charge: 2 NH3+ o 2(-) charge: 2 COO- o Zwitterion form e = Add OH- until DH 9.82 (net charge: -1) o 50% of a-NH3+ of Asp donates a proton and becomes NH2 o 1(+) charge: NH3+ o 1(0) charge: NH2 2(-) charge: 2 COO- . add OH- until pH 10.53 (net charge: -2) o 50% of NH3+ of Lys donates a proton becoming NH2 o 2(0) charge: 2 NH2 o 2(-) charge: 2 COO- © IpH o 100%0netcharge o Zwitterion form o Between pH 3.86 and 9.82 0° 63.86 + 9.82/2 = 6.84 YES (Y: Tyr, E: Glu, S: Ser)=tyrosyl-glutamyl serine Tyr pKa: a-NH3+=9.11, RH= 10.07 Glu pKa: y-COOH= 4.25 Ser pKa: a-COOH= 2.21 (primary alcohol remains uncharged at any pH) * pHO-l (net charge: +1) o 1 (+) charge: NH3+ o 3 (0) charge: R-OH, 2 COOH e Add OH- until pH 2.21 (net charge: 0) o 50% COOH of Ser donates a proton becoming COO- o 1 (+) charge: NH3+ o 2 (0) charge: R-OH, COOH o | (-) charge: COO- of Ser e Add more OH- until pH 4.25 (net charge: -1) o Y-COOH of Glu donates a proton and becomes COO- o 1 (+) charge: NH3+ o 1 (0) charge: R-OH o 2(-) charge: 2COO- e ~=Add OH- until pH 9.11 (net charge: -2) o 50 %of a-NH3+ of Tyr donates a proton and becomes NH2 o 2 (0) charge: ROH, NH2 o 2(-) charge: 2COO- e Add OH- until pH10.07 (net charge: -3) o 50% of Phenolic group (benzene-OH) donates a proton becoming O- o 1 (0) charge: NH2 o 3/(-) charge: 2 COO, R-OH e = Higher pH will still have a net charge of -3 (but 100% rather than 50%) ° pH o 2.21+4.25/2= 3.23 o Exact pH 3.23 when the peptide YES is 100% zero net charge/zwitterion e@ pH <IpH = ionic form of the peptide is positively (+) charged ARTICONA — 2FMT