Biochem Amino Acids

• Amino acids are the building blocks of proteins. The versatility of proteins comes from their primary sequence of amino acids. The sequence of amino acids dictates the final protein structure and function.
• The common structure of amino acids is the amino group, carboxylic group, Hydrogen (proton), and R group.
• Can a single polypeptide chain achieve quaternary structure?

  No- quaternary requires multiple polypeptide chains
• The only amino acid that does not have a chiral center is glycine.
• The D isomer favors carbohydrates, and the L isomer favors proteins. If the D isomer spins clockwise, then the L isomer spins counterclockwise. The D isomer has the hydroxyl group located on the right; therefore, the L isomer has the hydroxyl group on the left.
• In dipolar form, the amino group is protonated and the carboxyl group is deprotonated. In ionized form, the amino group is protonated, but the carboxyl group is not dissociated.
• Amino acids with non polar, aliphatic R groups:

  1. glycine
  2. alanine
  3. proline
  4. valine
  5. leucine
  6. isoleucine
  7. methionine
• Hydrophobic amino acids:

  1. valine
  2. leucine
  3. isoleucine
  4. methionine
  5. proline
  6. phenyalanine
  7. tryptophan
• Amino acids with polar, uncharged R groups:

  1. serine
  2. threonine
  3. cysteine
  4. asparagine
  5. glutamine
• Hydrophilic amino acids:

  1. Aspartic acid
  2. Glutamic acid
  3. Glycine
  4. Proline
  5. Serine
• Glutamate is an integral component of amino acid metabolism and nucleic acid base synthesis: 2 cysteine molecules can form disulfide linkages that bridge two separate polypeptide chains or sections within the same polypeptide.
• Amino Acids with Aromatic R groups:

  1. Phenylalanine
  2. Tyrosine
  3. Tryptophan
• Nonessential amino acids:

  1. alanine
  2. arginine
  3. asparagine
  4. aspartate
  5. cysteine
  6. glutamate
  7. glutamine
  8. glycine
  9. proline
  10. serine
  11. tyrosine
• Essential amino acids:

  1. histidine
  2. isoleucine
  3. leucine
  4. lysine
  5. methionine
  6. phenylalanine
  7. threonine
  8. tryptophan
  9. valine
• Positively charged R groups:

  1. lysine
  2. arginine
  3. histidine

  these are considered basic amino acids
• Positively charged amino acids are involved in electrostatic interactions, mainly on the surface of soluble proteins.
• Lysine is integral in catalytic sites where they bind the coenzymes biotin and lipoic acid.
• Histidine acts as a buffer.
• Negatively charge R groups:

  1. aspartate
  2. glutamate

  these are considered acidic amino acids
• There are 7 dissociable, and 13 nondissociable amino acids.
• When the pH is equal to 0, the amino acid will be protonated and very acidic.
• At a low pH all amino acids are positively charged.
• The zwitterionic form of an amino acid contains both the positive and negative charge.
• If you start from a lower pH, all amino acids eventually become mostly negative.
• The isoelectric point is the pH where the amino acid has no net charge.

  When pH < pI, there is a net positive charge.

  When pH = pI, there is a net neutral charge.

  When pH > pI, there is a net negative charge.

  To find the pI of an acidic amino acid, you will average the carboxyl pKa.

  To find the pI of a basic amino acid, you will average the amino pKa.

  To find the pI of a neutral amino acid, you will average only 2 pKa's
• Acidic amino acids will have a pI around 3.
• Basic amino acids will have a pI around 9.
• Neutral amino acids will have a pI around 6.
• Protein Purification Techniques:

  1. cells must be lysed- proteins released
  2. fractionated- centrifuged
  3. purified by manipulating the intrinsic properties of protein samples (solubility, charge, size, binding affinity)
• Salting Out:

  1. when you have proteins in solution, it is hydrated in an aqueous solvation cell
  2. add a competitor (ammonium sulfate) and it will remove the water molecules
  3. the proteins want to interact with themselves, and they will precipitate out if they huddle together enough
  4. some proteins require only a little bit of salt to reach the threshold if they are less soluble
  5. you will get a pellet with a lot of salt in it
• Dialysis:

  1. take a sample and put it in a semipermeable bag with 2 clips- lighter on top, heavier on bottom
  2. 10-12k molecular weight (everything less than 12k will go through, everything more will stay in)
  3. the objective is to get rid of the salt
• Ion exchange chromatography (cation exchange):

  1. positively charged protein binds to negatively charged bead
  2. negatively charged protein flows through
  3. pack the column with the stationary phase (something with a sulfate group)