Biochem Primary structure, Sequencing, Mass spectrometry and Sequence Alignments Reading: Chapter 5

Lec 11

Nature of Protein Sequences

The number of possible sequences is - small or large?

Astronomically large

Nature of Protein Sequences

What is the probability that two proteins will by chance have similar sequences

What is the result of this → What does sequence similarity imply?

the probability that two proteins will by chance have the similar sequences is negligible

Therefore, sequence similarity implies evolutionary relatedness

Nature of Protein Sequences

What are proteins with similar sequences described as?

What are the other two?

Proteins with similar sequences are describes as homologous

• orthologous proteins are from different species (ancestral gene)

• Paralogous proteins are from the same species (gene duplication)

Sequence Alignments

What do com[puter programs optimally align sequences in

computer programs optimally align sequences in databases - like BLAST or Basic Local Alignment Search Tool

Nature of Protein Sequences

What do mutations represent?

what is score alignment based on

G

What can the alignment be?

Mutations represent change at one or more sites in the amino acid sequence of a protein

Score alignment is based on matching or similar amino acids

Also put in gaps which bring penalties

Alignment can be local or global

Phylogeny of Cytochrome c

What is the number of amino acids differences between two cytochrome C sequence proportional to?

What can this observation be use for?

What is this the basis for?

the number of amino acids differences between two cytochrome c sequences is proportional to the phylogenetic difference between the species from which they are derived

This observation can be used to build phylogenetic trees of proteins

This is the basis for studies of molecular evolution

What do phylogenetic trees depict

what do the numbers represent

evolutionary relationship among eukaryotic organisms as determined by their similarity of their cytochrome c sequences

the numbers are amnio acid changes between hypothetical branch points and to the extent species at the tips

Protein Structure and Function

Structure depends on ______ and function depends on _____

structure depends on sequences and function depends on structure

not all proteins of similar structure share a common function -true or false

explain

true

some proteins share similar structural features but carry out different functions

proteins with different structure can carry out similar functions

How do proteins get structure?

peptide bond

links amino acids

partial double bond character

• coplanar O, C, N

• limited rotations

NH acts as a donor and O as an acceptor for H bonds

Phi (Φ) and Psi (Ψ) Angles

What are the degrees of freedom?

Positive values are what rotations

Only two degrees of freedom for peptide backbone (dihedral angles)

1. Angle about the Cα-N bond is denoted φ (phi)

2. Angle about the Cα-C bond is denoted ψ (psi)

positive values are clockwise rotations as view from Alpha and start at 0 degrees

Restriction of Phi (Φ) and Psi (Ψ) Angles

What values are forbidden and why?

Some values are forbidden due to steric crowding

φ=0°, ψ=180°

φ = 180°, ψ = 0°

φ=0°, ψ=0°

Display φ and ψ on a Ramachandran Plot

What are the colored regions?

What is prolines fixed phi angel?

What can particular angles give? (what type of structures)

the colored regions are allowed

Proline has a fixed phi angle of -60

particular angles give secondary structures

Protein Folding – 2°, 3°, and 4°

What do weak interactions do?

• VDW are…

• HB form….

• IB occur…

• HPI occur..

Weak interactions stabilize proteins

• can Der Waals are ubiquitous

• hydrogen bonds form wherever possible iconic bonds occur on protein surface

• hydrophobic interaction occur in protein interior/core and drive folding

Protein Folding – 2°, 3°, and 4°

the most stable structure has what?

What do H bonding involves?

What should happen to the polarity of H bonding groups if they are in the hydrophobic core?

the most stables structure has large no. of interactions

hydrogen bonding involves backbone CO and NH, plus some side chain groups

polarity of H-bonding groups must be neutralized if they are in the hydrophobic core. this occurs via secondary structures

Secondary and Supersecondary Structures

Secondary structures:

• helices (particularly alpha)

• beta sheets

• beta turns

• random coil

Supersecondary structures:

• coiled coils

• beta hairpin

• beta-alpha-beta

• helix-turn-helix, greek keys, alpha-alpha hairpin, EF hand etc.

What do h bonds do in the helix structure

they stabilize the helix structure

The α-Helix

how many residues per turns

φ= ψ=

what is the diameter

What is the helix pitch or rise

3.6 residues per turn

φ=−60°,ψ=−45 to -50°

diameter of 6 A (without sidechains)

The α-Helix

What is the helix pitch or rise

What are the helix lengths

what is the average number of residues per helix

helix pitch or rise of 5.4 A (1.5 A per res.)

different helix lengths

average of 10 reissues per helix

The α-Helix

is it right or left handed and why

are left handed or right handed helices shorter

almost always right handed due to close approach of side chains of amino acids (for L-amino acids, the left hand alpha-helix has a close approach between the carbonyl oxygen and the Beta-carbon)

left hand helices are shorter and contain glycine