Topic 1 Amino Acids, peptides and proteins
We will restrict our discussion to α-amino acids as these are the ones that are found in biology.
So why are they called α?
We talk about the Cα, Cβ and Cγ atoms of amino acid side chains (later)
In Biology, one tends to think of the basic building block as
Proteins are only made with (S)-amino acids (except cysteine which is R) but they are called L-amino acids. This derives from the Fischer projection.
In a Fischer projection put the molecule flat onto paper like so
Racemisation
Although proteins are made with L – amino acids, the other hand (D or R) are common in drugs and poisons.
These are made by a multitude of biosynthetic pathways.
Direct racemisation is possible by α – proton abstraction. This is chemically difficult (pKa > 15) but enzymatically facile
Acid or base?
We already know that NH2 groups are basic (they like to accept a proton) and CO2H groups are acidic (they like to donate a proton).
At what pH is the CO2H group protonated?
(below 3)
At what pH is the NH2 protonated ?
(below 10)
These numbers are called the pKa.
What happens at pH 7?
The acid is deprotonated and amine is protonated, the amino acid has this structure
This is called a zwitterion
The nature of R
There are twenty amino acids and 19 of them have this simple building block only the R-group varies.
Acidic, charged
glutamic acid, Glu, E,
- CH2 – CH2 – CO2H , pKa 4.3
aspartic acid, Asp, D,
- CH2 – CO2H , pKa 3.7
Basic, charged
arginine, Arg, R, -
CH2 – CH2 – CH2 – NH - C = NH(NH2), pKa 12.5
lysine, Lys, K,-
CH2 - CH2 - CH2 – CH2 – NH2, pKa 10.3
Basic, uncharged
Histidine, His, H
pKa 6.0
Hydrophobic
alanine, Ala, A - CH3
valine, Val, V , - CH – (CH3)2
leucine, Leu, L, - CH2 - CH – (CH3)2
isoleucine, Ile, I , - CH –CH3 (CH2-CH3)
methione, met, M -CH2–CH2–S-CH3
phenylalanine, Phe, F
tryptophan, Trp, W
Structural (hydrophobic)
glycine, Gly,G - H
proline, Pro, P
Polar
serine, Ser, S,
- CH2 – OH , pKa 9.2
threonine, Thr, T
- CH – OH (CH3) , pKa 9.2
cysteine, Cys, C
- CH2 – SH , pKa 8.3
asparagine, Asn, N
glutamine, Gln, Q
tyrosine, Tyr, Y
pKa 10.1
Charged and polar amino acids like to be in contact with charged polar amino acids or in a polar environment, hydrophobic in an apolar environment.
What chemistry can we do with amino acids?
Acylation
What is the nucleophile?
What makes it a good nucleophile and why?
What sort of electrophile might you use?
Ninhydrin detection of amino acids
Making amino acids and the peptide bond
Synthesis of amino acids
Simplest is treatment of α-halo acid with ammonia. The bromo acids are easy to get hold of. The competing SN2 reaction with the amino acid is slow and excess ammonia is used.
(Jones Organic Chemistry)
A favourite method of making amino acids is to use the Strecker synthesis which is based on the use of cyanide
(Jones Organic Chemistry)
Both of these methods would produce a racemic mixture (both D & L). We want only one, stereochemistry is very important. How can we separate enantiomers? (Called resolution)
Enantiomers have identical chemical properties, diastereomers have different chemical properties. We need to make diastereomers. So we react racemic amino acids with an other optically pure material, the result is two diastereomers which can separated.
More commonly is to use enzymes which will react with only D or L or which make only D or L. In fact genetic engineering can be used to make bacteria which will synthesis a particular amino acid. (see Jones, p1280)
What kind of chemistry can the side chains do?
Nucleophile
Lys Cys > Ser (Thr) > Tyr >> His > Asp(Glu)
Why are Asn and Gln not nucleophiles?
Lewis base
Met (S-adenosyl methionine)
Acid and Bases
His, Glu, Asp, Lys, Arg and very rarely Tyr
Cys has special chemistry it is able to form a disulphide bond
(Jones, Organic Chemistry)
Amino acids react together to form peptides and proteins.
As a definition, a peptide is about 50 residues and a protein is more.
The driving force is the elimination of water. This is very favourable. The peptide bond is very kinetically stable, proteins do not decompose easily without intervention.
Where does the stability in the peptide bond come from?
Consider the hybridisation of the atoms at the peptide bond
Atom 1 is an sp3 carbon
Atom 2 is an sp2 carbon
Atom 3 is a nitrogen
Atom 4 is an sp3 carbon
Look closely at the nitrogen
This is known as resonance
In effect the bond between atom 2 and atom 3 has double bond character.
The amide nitrogen has sp2 hydridisation
In Sp2 hybridised nitrogen, the lone pair is unavailable (involved in resonance). The amide nitrogen has no nucleophilic chemistry at all.
Peptides have a directional sense for instance
These are different compounds.
We always write peptides (and proteins) as a sequence of letters. We start from the N-terminus. Left is SC and right is CS. The reason we start from the N-terminus is because so does biology. (Even short peptides can have dramatic biological effects. Nutrasweet and oxytocin are good examples of such peptides.)
We talk about protein sequence or amino acid sequence. It is also known as primary structure of a protein.
How do we determine the primary structure (or sequence)?
Sanger, a chemist whom the Sanger sequencing centre is named after, developed a chemical method to decompose peptides and identify the first residue. However, it is the Edman degradation which allows one to sequence proteins.
(Jones, Organic Chemistry)
With Edman one can get about 20 to 30 residues along before impurities mask the signal in noise. A really ideal system can reach 50 but that is very rare. So how are longer proteins sequenced?
They are cut into smaller bits and each fragments. In the real world everyone uses enzymes, mainly Trypsin which cuts after Lys and Arg.
Trypsin belongs to a family of enzymes known as serine proteases.
These are great enzymes which do fantastic chemistry
A few words about peptide synthesis
In biology proteins are made by reading the genetic code, they are assembled one by one starting at the N-terminus.
Imagine making a nine – mer. If each step of elongation was 95% effective, even for a 9mer the overall yield is 63%. Think of the steps you would need to do, remember SC and CS are different! To overcome this, one uses protecting groups. Proteins are always synthesised on a solid phase (higher yield) but are made from the C-terminus. Convergent synthesis is required to make larger peptides, proteins of 100’s of amino acids have been made.
Secondary and tertiary structure
We talk about the peptide chain and in fact some chemistry text books used to show peptides (and proteins) as beads on a string. However, proteins have a structure.
This in part arises from the restricted rotation around the peptide bond (Why?)
In addition, there are orientations between peptides which are more and less favourable
Proteins fold in strands, helices, turns or coils. These come together (folding) to make the following secondary structural units.
Protein structure relies on interactions between amino acids.
These are
Covalent (disulphide) bonds
Hydrophobic / hydrophobic interactions (VDW)
Hydrogen bonds (2.5Å to 3.5Å) optimum 2.8Å and co-linear
Salt bridges 2.5Å to 4.0Å
As they form these units, these fold into domains and then into a full structure.
What sort of things contribute to protein folding
FAVOURABLE UNFAVOURABLE
DH
H-bonds made H-bonds broken
Ionic bonds
Van der Waals
DS
"Hydrophobic Effect" 1 Conformation
Proteins are only just folded in energetic terms.
Why is this an advantage?
A monomer is a single continuous piece of sequence
Often though the active protein is made up from more than one monomer. On can see the same protein above in its natural and active dimer form.
The structure of proteins is determined by a procedure known as X-ray crystallography.