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Amino Acid and Protein

Essential amino acids

  • Essential amino acids cannot be synthesized in our body.

Stereochemistry of amino acids

Chirality and enantiomers

  • Many amino acids exist as enantiomers (chiral).

  • Enantiomers rotate plane-polarized light, measurable with a polarimeter:

  • d / (+): rotates light to the right (dextrorotatory, clockwise)

  • l / (-): rotates light to the left (levorotatory, counterclockwise)

D/L configuration (relative to glyceraldehyde)

  • L- is similar to L-glyceraldehyde:

  • COOH top, Meth left, H back, NH\(_2\) front

  • D- is similar to D-glyceraldehyde:

  • COOH top, Meth right, H back, NH\(_2\) front

Special cases and biological usage

  • Glycine is achiral (no L-glycine or D-glycine).
  • Bacteria can use D-amino acids.
  • Humans use L-amino acids.
  • Thalidomide (pain killer) caused issues because it is racemic and toxic.

Amino acid classes (by side chain properties)

Nonpolar, aliphatic (no double bond): hydrophobic

  • Glycine (Gly)
  • Alanine (Ala)
  • Proline (Pro)
  • Valine (Val)
  • Leucine (Leu)
  • Isoleucine (Ile)

Proline note :

  • Proline is the only one whose side chain forms a bond back to the backbone and is often used when a chain needs sharp turns.

Polar, aliphatic, uncharged (with hydroxyl, carboxyl, or amide)

  • Asparagine (Asn)
  • Glutamine (Gln)
  • Serine (Ser)
  • Threonine (Thr)

Aromatic

  • Phenylalanine (Phe)
  • Tyrosine (Tyr)
  • Tryptophan (Trp)

Aromatic absorption :

  • Phe, Tyr, Trp absorb UV light at 280 nm.

Beer–Lambert Law

\[ A(\text{absorbance})=\varepsilon (\text{absorb. coeff}),l(\text{length of fluid}),c(\text{conc.}) \]

Acidic amino acids

  • Aspartate (Asp)
  • Glutamate (Glu)

Feature:

  • carboxyl group in the side chain

Basic amino acids

  • Arginine (Arg)
  • Lysine (Lys)
  • Histidine (His)

Note :

  • For His, it is charged if the solution condition is optimal.

Feature:

  • amino group in the side chain

Sulfur-containing amino acids

  • Methionine (Met)
  • Cysteine (Cys)

Note:

  • Cys can form disulfide bonds.

Ionization states and amphoteric behavior

  • Low pH: cation, pK\(_{a1}\) approx 2
  • Neutral pH: zwitterion
  • High pH: anion, pK\(_{a2}\) approx 10

Amino acids are amphoteric: they possess both acidic and basic groups.

Henderson–Hasselbalch equation

\[ \text{pH}=\text{p}K_a+\log_{10}\frac{[\text{A}^-]}{[\text{HA}]} \]
  • \(K_a\) is the dissociation constant.

Isoelectric point (pI) and buffering

  • Isoelectric point (pI): average of the two pK\(_a\) values that are the closest; amino acids are in the zwitterion state there .
  • Amino acids are weak acids and weak bases, so they can buffer.
  • Buffering capacity is best at its pK\(_a\) (50% can donate/accept a proton), and worst at the zwitterion point .

Protein structure

1° (primary) structure

  • polypeptide chain connected by peptide bonds
  • peptide bond formation was facilitated with 28S rRNA

Polyampholytic behavior:

  • proteins can have multiple ionizable groups contributing to overall charge behavior.

Isoelectric point (protein version):

  • pI is the average of pK\(_a\) of all ionizable groups.

2° (secondary) structure

  • hydrogen bonds form between peptide bonds

Alpha-helix :

  • C=O … H–N (in four amino acids)
  • R group points out and H points out, clockwise

Beta-pleated sheet :

  • two or more strands aligned laterally
  • may be inter/intra chain
  • may be parallel or anti-parallel
  • R groups alternate to the two sides of the sheet

3° (tertiary) structure

  • involves functional group interactions, e.g.:

  • hydrogen bonds

  • hydrophobic interactions
  • disulfide bridges
  • ionic bonds
  • achieves the state with the lowest internal energy

4° (quaternary) structure

  • interactions within different subunits

Purification

Chromatography

Isolates proteins from mixtures.

Size separation

  • molecules larger than the bead pore size pass through quicker

Ion exchange

Example :

  • cation exchange: beads with negatively charged ions; cations will not pass through and anions will pass through (want protein to bind to beads)

Affinity chromatography

  • bead is conjugated to a ligand that binds the protein
  • the protein is washed off the complex