Protein Structure

Dr. Tapan Kr. Dutta

Panskura Banamali College
From: Protein Data Bank PDB ID: 1B0E
Kalus, W., Zweckstetter, M., Renner, C., Sanchez, Y., Georgescu, J., Grol, M., Demuth, D., Schumacher, R., Dony, C., Lang, K., Holak, T. A.: structure of the IGF-binding
domain of the insulin-like growth factor-binding protein-5 (IGFBP-5): implications for IGF and IGF-I receptor interactions. EMBO J 17 pp. 6558 (2010)
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Functions
Diverse functions related to structure
Structural components of cells
Motor proteins
Enzymes
Antibodies
Hormones
Hemoglobin/myoglobin
Transport proteins in blood
Importance of Proteins

Muscle structure depends on protein-

protein interactions
Transport across membranes involves
protein-solute interactions
Nerve activity requires transmitter
substance-protein interactions
Immune protection requires antibody-
antigen interactions
Levels of Protein Structure
Charged/polar R-groups generally
map to surfaces on soluble
proteins
Non-polar R-groups tend to be buried
in the cores of soluble proteins

Myoglobin

Blue = non-polar
R-group

Red = Heme
Membrane proteins have adapted
to hydrophobic environments
Some R-groups can be ionized
The Henderson-
Hasselbalch
equation allows
calculation of the
ratio of a weak acid
and its conjugate
base at any pH

( )
protonated
unprotonated
Structure

Amino acids
Amino group
(NH2)
Carboxyl
group (COOH)
From: Elliott, WH. Elliott, DC. (1997) Biochemistry and
Molecular Biology. Oxford: Oxford University Press. p23
Stereochemistry
Configuration
of amino acids in proteins
The CORN Law
Amino acids
20 amino acids make
up protein
8 essential amino
acids
9 in infant (histidine)
From: Elliott, WH. Elliott, DC. (1997) Biochemistry
and Molecular Biology. Oxford: Oxford University
Dipolar
+ve end (NH3+)
Press. p23

-ve end (COO-)

Overview

Primary Structure
Secondary Structure
Tertiary Structure
Quaternary Structure
 Peptide Bond

The peptide bond influences all aspects of protein structure and

function.
Key features:
1. Planar
2. Fairly rigid, due to partial double bond character.
3. Almost always in trans configuration.
4. Polar. Can form at least two hydrogen bonds.
5. Places restrictions on the conformation of the polypeptide chain.
Primary Structure
Polypeptide chains
Amino Acids
Largest polypeptide
chain approx has
5000AA but most have
less than 2000AA
Amino Acid Basic
Structure H2N-CH-
National Genome Research Institute
genome.gov
COOH
Arrangement of the 20
amino acids in the
polypeptide is the
amino acid sequence
which composes the
primary structure of the
protein
20 Amino Acids
Nonpolar,
hydrophobic

Polar, uncharged

Polar, charged

http://www.people.virginia.edu/~rjh9u/aminacid.html
Amino Acid Classification

aliphatic
G P
beta-branched
CS-S A
CH S N
V
L
I T D
Q
E
negative
M K
F Y H R
aromatic W
positive
hydrophobic charged
polar
Bioinformatics Methods II, Spring 2003

A Venn diagram showing the relationship of the 20

naturally occurring amino acids to a selection of physio-
chemical properties thought to be important in the
determination of protein structure.
Protein structure - bonding

5 bonds or forces determine

structure
Peptide bond
Hydrogen bond

Disulfide bond

Ionic bond

Hydrophobic force
Peptide Bond

Resonance
C-N bond length of the peptide is
10% shorter than that found in
usual C-N amine bonds
Peptide bond planer
, angle around peptide bond,
00 for cis, 1800 for trans
Peptide Bond

http://cmgm.stanford.edu/biochem/biochem201/Slides/
Peptide bond

Peptide bond joins

amino acids
Bond at both ends
Increases range of
possible proteins
2 peptides can
result from bonding
of 2 amino acids
1.0 x 1026 peptides
can be formed from
20 amino acids From: Elliott, WH. Elliott, DC. (1997) Biochemistry
and Molecular Biology. Oxford: Oxford University
Press. p23
Bond Formation

Linking two amino

acids together

Definitions (N-
terminal, C-terminal,
polypeptide
backbone, amino
acid residue, side
chains) http://web.mit.edu/esgbio/www/lm/proteins/peptidebond.html
Primary Structure

What is a native protein?

Protein conformation &
problem of protein folding
Hydrophobic, hydrophilic
Charge
Chaperones
Special Purpose Amino Acids
Proline Cysteine
Primary protein structure Primary structure of insulin

Linear sequence of
amino acids forms
primary structure
Sequence essential for
proper physiological
function

Bettelheim & March (1990) Introduction

to Organic & Biochemistry
(International Edition) Philadelphia:
Saunders College Publishing, p299
 Sickle cell anemia

Replacement of
single glutamine
with valine in one
polypeptide chain
of hemoglobin
alters structure
and function
Bettelheim & March (1990) Introduction to Organic & Biochemistry
(International Edition) Philadelphia: Saunders College Publishing, p301
The peptide bond is planar

This resonance
restricts the number
of conformations in
proteins -- main
chain rotations are
restricted to f and y.
 The a-helix
In the a-helix, the
carbonyl oxygen of residue
i forms a hydrogen bond
with the amide of residue
i+4.

Although each hydrogen

bond is relatively weak in
isolation, the sum of the
hydrogen bonds in a helix
makes it quite stable.

The propensity of a
peptide for forming an a-
helix also depends on its
sequence.
 Tight Turns
There are different types of tight turns (Chou 2000)
depending upon the number of atoms forming the
turn. These are as follows:

Delta-turn - It is the smallest tight turn which involves

only two amino acid residues and the intraturn
hydrogen bond for a delta-turn is formed between the
backbone NH(i) and the backbone CO(i+1).
Gamma-turn - It involves three amino acid residues
and the intraturn hydrogen bond for a gamma-turn is
formed between the backbone CO(i) and the backbone
NH(i+2).
 Tight Turns
Beta-turn - A beta-turn involves four amino acid
residues and may or may not be stabilized by the
intraturn hydrogen bond between the backbone
CO(i) and the backbone NH(i+3).
Alpha-turn - An alpha-turn involves five amino
acid residues where the distance between the
Calpha(i) and the Calpha(i+4) is less than 7 and
the pentapeptide chain is not in a helical
conformation.
Pi-turn - It is the largest tight turn which involves
six amino acid residues.
 -turn

A -turn is a region of the protein

involving four consecutive residues
where the polypeptide chain folds
back on itself by nearly 180 degrees
The b-sheet
In a b-sheet,
carbonyl oxygens and
amides form hydrogen
bonds.
These secondary
structures can be
either antiparallel (as
shown) or parallel and
need not be planar (as
shown) but can be
twisted.

The propensity of a
peptide for forming b-
sheet also depends on
its sequence.
 b turns

b-turns allow the protein backbone to make abrupt turns.

Again, the propensity of a peptide for forming b-turns depends
on its sequence.
b turns
b turns
Ramachandran
plot -- shows f
and y angles
for secondary
structures
Tertiary structures are quite varied
Secondary protein structure

Peptidechains fold into secondary

structures:
a - helix
b - pleated sheet
Random coil
Secondary protein structure
Secondary protein structure
Secondary protein structure
Protein Secondary Structure

Introduction
Peptide bond geometry
Ramachandran plot
Structures
Regular local structures formed
by single strands of peptide
chain due to constraints on
backbone conformation
Ramachandran Plot

http://hykim.chungbuk.ac.kr/lectures/biochem/4-5/fig6-9(L).jpg
Alpha Helix

http://cmgm.stanford.edu/biochem/biochem201/Slides/
Alpha Helix
Left-handed Right-handed

http://www.rtc.riken.go.jp
Alpha Structure Features

3.6 residues per turn

5.4 angstroms in length per turn
carboxyl group of residue i
hydrogen bonds to amino group
of residue i+4
 Helix Structures
H Bond R/t A/t
Alpha -57.8 -47 i, i + 4 3.6 13

3-10 Helix -49 -26 i, i + 3 3.0 10

Pi Helix -57 -80 i , i + 5 4.4 16

http://broccoli.mfn.ki.se
More Helix Structures

Type comments

Collagen -51 153 Fibrous proteins

Three left handed helicies
(GlyXY)n, X Y = Pro / Lys

Type II helices -79 150 left-handed helicies formed

by polyglycine
Beta Sheet

http://www.rothamsted.bbsrc.ac.uk/notebook/courses/guide/images/sheet.gif
Beta Sheet Features

Sheets can be made up of any

number of strands
Orientation and hydrogen bonding
pattern of strands gives rise to flat or
twisted sheets
Parallel sheets buried inside, while
Antiparallel sheets occurs on the
surface
 More Beta Structures
Beta Bulge chymotrypsin
(1CHG.PDB) involving
residues 33 and 41-42

Anti parallel
Beta Twist pancreatic trypsin
inhibitor (5PTI) 0 to 30 degrees per
residue
Distortion of tetrahedral N atom
http://broccoli.mfn.ki
 Beta turns

i + 1 Pro
i + 2 Pro or Gly
i + 3 Gly

http://rayl0.bio.uci.edu/~mjhsieh/sstour/image/betaturn.png
Interactions

Covalent bonds
Disulphide bond (2.2 0A) between two Cys
residues
Non-covalent bonds
Long range electrostatic interaction
Short range (4 0A) van der Waals interaction
Hydrogen bond (3 0A)
a - helix
Shape
maintained by
hydrogen bonds
between C=O
and N-H groups
in backbone
R groups directed
outward from coil
From: Elliott, WH. Elliott, DC. (1997) Biochemistry and Molecular
Biology. Oxford: Oxford University Press. p28
b - pleated sheet

Structure
maintained by
hydrogen bonds
between C=O and
N-H groups in
backbone
R groups directed
above and below
backbone
From: Elliott, WH. Elliott, DC. (1997) Biochemistry and Molecular
Biology. Oxford: Oxford University Press. p29
 Not really random
structure, just non-
Random coil repeating
Random coil has
fixed structure within
a given protein
Commonly called
connecting loop
region
Structure determined
by bonding of side
From: Elliott, WH. Elliott, DC. (1997) Biochemistry and Molecular
Biology. Oxford: Oxford University Press. p27 chains (i.e. not
necessarily hydrogen
bonds)
Tertiary protein structure

Secondary structures fold and pack

together to form tertiary structure
Usually globular shape
Tertiary structure stabilised by bonds
between R groups (i.e. sidechains)
Tertiary structure - H bond
H bonds weak
allowing to be
broken and
reformed easily
Allows structural
Hydrogen bond
change
produces
functional
molecules
Tertiary Protein Structure

Defines the three dimensional

conformation of an entire peptide chain in
space

Determined by the primary structure

Modular in nature
Aspects which determine tertiary
structure

Covalent disulfide bonds from between

closely aligned cysteine residues form the
unique Amino Acid cystine.
Nearly all of the polar, hydrophilic R groups
are located in the surface, where they may
interact with water
The nonpolar, hydropobic R groups are
usually located inside the molecule
Motifs and Domains
Motif a small structural domain
that can be recognized in a variety of
proteins
Domain Portion of a protein that
has a tertiary structure of its own. In
larger proteins each domain is
connected to other domains by short
flexible regions of polypeptide.
 Modular Nature of Protiens
Epidermal Growth
Factor (EGF) domain
is a module present
in several different
proteins illustrated
here in orange.
Each color
represents a
different domain
Domain Shuffling
Occurs in evolution
New proteins arise by joining of
preexisting protein domain or modules.
Tertiary structure - disulfide
bond

Covalent bond
between sulfur
atoms on two
cysteine amino
acids
From: Elliott, WH. Elliott, DC. (1997) Biochemistry
and Molecular Biology. Oxford: Oxford University
Press. p32
Tertiary structure - ionic bond

Ions on R
groups form salt
bridges through
ionic bonds

From: Summerlin, LR. (1981) Chemistry for the Life Sciences. New
York: Random House, p459
Tertiary structure - hydrophobic
forces
Close attraction of non-
polar R groups through
dispersion forces
Very weak but collective
interactions over large
area stabilise structure
Repel polar and charged
molecules/particles Bettelheim & March (1990) Introduction to Organic & Biochemistry
(International Edition) Philadelphia: Saunders College Publishing, p302
Quaternary protein structure
Arrangement of
multiple tertiary
structures into single
functional complex
Allows for changes in
structure/function in
response to chemical
stimuli From: Elliott, WH. Elliott, DC. (1997) Biochemistry and
Molecular Biology. Oxford: Oxford University Press. p27
Quaternary Structure
non-linear
3 dimensional
global, and across distinct
amino acid polymers
formed by hydrogen
bonding, covalent
bonding, hydrophobic
packing and hydrophilic
exposure
favorable, functional
structures occur frequently
and have been categorized
 Quaternary Structure
Not all proteins have a
quaternary structure
A composite of multiple
poly-peptide chains is
called an oligomer or
multimeric
Hemoglobin is an
example of a tetramer
Globular vs. Fibrous
Protein Folding

Proteinfolding constitutes the

process by which a poly-peptide
chain reduces its free energy by
taking a secondary, tertiary, and
possibly a quaternary structure
Thermodynamics
Proteins follow
spontaneous
reactions to reach
the conformation
of lowest free
energy
Reaction
spontaneity is
modeled by the
equation G=
H-TS
 Molecular Visualization
Goal: Clear
visualization of
molecular structure
Different visualization
modes elucidate
different molecular
properties
Some representations
include Ribbons, Space
Fill and Backbone
Class/Motif
class = secondary structure
composition,
e.g. all a, all b, segregated
a+b, mixed a/b
motif = small, specific
combinations of secondary
structure elements,
e.g. b-a-b loop
both subset of
fold/architecture/domains
Fold/Architecture/Domains
fold = architecture = the overall
shape and orientation of the
secondary structures, ignoring
connectivity between the structures,
e.g. a/b barrel, TIM barrel
domain = the
functional property
of such a fold or
architecture,
e.g. binding, cleaving, spanning sites
subset of topology/fold
families/superfamilies
Topology/Fold families/Superfamilies

topology = the overall shape

and connectivity of the folds
and domains
fold families = categorization
that takes into account
topology and previous subsets
as well as empirical/biological
properties, e.g. flavodoxin
superfamilies = in addition to
CLASS: a+b
fold families, includes
FOLD: sandwich evolutionary/ancestral
FOLD FAMILY: flavodoxin
properties
CS/Math/Physics of Protein Structure

Experimental Determination
and Analysis
Computational Determination
and Analysis
NMR Spectroscopy
protein in aqueous solution,
motile and tumbles/vibrates
with thermal motion
NMR detects chemical shifts
of atomic nuclei with non-
determining constraints zero spin, shifts due to
electronic environment
nearby
determine distances between
specific pairs of atoms based
on shifts, constraints
use constraints and
biochemical knowledge of
the protein to determine an
using constraints to determine
secondary structure
ensemble of models