Membrane Proteins

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Membrane Proteins Lecture 7 BINF 5513
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Proteins can be divided into 3 main classes (1)
Fibrous protein, form vast aggregates structure
is usually highly hydrogen-bonded, highly regular
and maintained mainly by interactions between
various chains. (2) Membrane proteins live in a
water-lacking membrane structures are highly
regular (like fibrous proteins) and highly
hydrogen-bonded. (3) Water-soluble globular
proteins. live in water structures are less
regular. Important role of hydrophobic effect.,
These three classes are distinguished by

1)living, environmental conditions,

2) structure-stabilizing interactions,
and

3) overall architecture of proteins.
Question Explain, please, how do
you understand these 3 points.
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Structure of a typical cell membrane
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Cell, Membrane - definition
Cells are structural units, compartment, that
make up plants and animals. One organism, like
the human, can have the same genetic material in
every cell. There are over 200 types of cells in
the human, that are different shapes, sizes and
carry out very different functions. Cell
membrane are the structure separating an cell
from its environment. The cell membrane is a
complex system that allows nutrients to enter the
cell and waste products to leave.
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Membrane Structure and Function
Membranes are vital  because they separate the
cell from the outside world.  They also separate
compartments inside the cell to protect important
processes and events.
The electron microscopic figure shows a typical
common structure which resembles a railroad track
with two dense lines separated by a clear space. 
why a railroad track ?
Membranes include lipids and protein molecules.
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The membrane include the phospholipids
polar hydrophilic head
lipids are both hydrophilic and hydrophobic
     
The role of lipids polar heads
pointing out to the water and
the hydrophobic part forming the
core of the membrane
two hydrocarbon tails.
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The First Membrane model In the 1930's-40's,
Danielli and Davson studied lipid bilayers over a
water surface. Lipids arranged themselves with
the polar heads facing outward and always formed
droplets (oil in water).    
However, if add proteins, the surface tension was
reduced and the membranes flattened out. 
Conclusion Membranes are composed of lipid
bilayers with proteins
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The next Membrane model In 1966, Lenard and
Singer noted that over 30 of membrane proteins
were twisted into an alpha helix.  Singer
studied phospholipid bilayers with the freeze
fracture techniques
a) rapid freezing of a cell ? b) the frozen
cells are cleaved along a fracture plane ?
c) an electron microscope You can see
protrusions or bumps ( colored red ) and
see
structures within the bumps. These are the
transmembrane proteins.
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Membrane Architecture Lipid bilayer. The lipids
hydrophilic polar heads pointing out and the
hydrophobic portion forming the
core. Transmembrane Proteins (half of the
membrane weight) pass through the bilayer.
The role of membrane proteins is to provide
transmembrane transport of various molecules and
signals.
The membrane
is a kind of insulator while its proteins act
as conductors.
These conductors are specific, each ensuring
transmembrane transport of molecules.
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The lipid bilayer gives the membranes its fluid
characteristics. 
At a low temperatures, the bilayer is

in a gel state and tightly packed. At
higher (body) temperatures,

the bilayer actually "melts' and

the interior is fluid
allowing
the lipid and
protein molecules
to move
around, rotate, exchange places.
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Two types of Membrane Proteins
I. Transmembrane proteins are amphipathic as
lipids, thus they have hydrophobic and
hydrophilic regions.
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II. The second types of proteins may be linked
only at the cytoplasmic surface or at the
external cell surface, these are called
"peripheral membrane proteins" . 
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• Membrane Architecture
• The proportion of proteins and lipids varies
  depending on the membrane.
• ?Nerve fibers, contains only 18 protein and 76
  lipid.
• ? Mitochondrial inner membrane contain 76
  protein and only 24 lipid.
• ? Plasma membranes of human red blood cells
  contain nearly equal amounts of proteins (44 )
  and lipids (43).

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Main Thermodynamic Principles of Membrane
Proteins First, most of the amino acid
side-chains of  transmembrane segments must be
non-polar. In fact, Residues with leucine,
isoleucine, alanine, valine) predominate in the
middle of the bilayer.
WHY?
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Second, the very polar CONH groups
(peptide bonds) of the polypeptide backbone of
transmembrane segments must participate in
hydrogen bonds (H-bonds) Why?

Hint polar group
usually have contact with water molecule, to
neutralize the charge the hydrogen bonds can be
formed because This H-bonding is most easily
accomplished with .
alpha-helices and with beta- sheets.
All
membrane proteins of known three-dimensional
structure adhere to these principles.

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How Membrane Proteins Are Assembled
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It is a complex process involving synthesis of
membrane proteins by ribosomes attached
transiently to a proteins translocon (sequence
of 20-22 hydrophobic amino acids, which located
within the cell membrane. This translocon
provides a transmembrane "tunnel" into which the
newly synthesized protein can be injected.

After the
synthesis is complete, the ribosome dissociates
from the translocon and the protein is released
into the membrane bilayer where it assumes (in an
unknown way) its final folded three-dimensional
structure.
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Non-constitutive membrane proteins incorporate
themselves into membranes of cells.
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Summary

The membrane proteins live within the
membrane where is practically no water and
therefore, the intra-membrane parts consist of
the regular secondary structure. Only few
membrane proteins have been solved. This is due
to their poor solubility in water and
difficulties of crystallization caused by their
tendency to disordered association.
Why?
All
Hydrogen bonds are busy with the formation of
secondary structure strands and helices, but
not contacts with water, as it is typically for
globular and fibrous proteins.
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from the Stephen White Laboratory at UC Irvine
http//blanco.biomol.uci.edu/Membrane_Proteins_xta
l.html
Unique Proteins in Database 111.
Number of
Coordinate Files in Database 211 Includes
proteins of same type from different species. For
example, photosynthetic reaction centers from R.
viridis and R. sphaeroides are considered
unique.Structures of mutagenized versions of
proteins already in the database are excluded as
unique. Proteins that differ only by substrate
bound or by physiological state are also excluded

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Bacteriorhodopsin (1C3W)
Pumps protons across the membrane
Transmembrane portion seven ?-helices that span
the bilayer. The membrane-exposed surface is
highly hydrophobic.Hydrophobic interactions hold
the proteins in the membrane The charged/polar
residues are at the solvent interfaces and also
line the internal hydrophilic channel.
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Hydrophobicity plot for the bacteriorhodopsin
molecule
Shows the hydrophobic regions
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• Bacteriorhodopsin contains retinal - a prosthetic
  group (?)

A tightly bound, specific non-polypeptide unit
required for the biological function of some
proteins
light

• retinal molecule that is covalently bound to a
  conserved lysine (Lys-216)

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Bacteriorhodopsin acts as a light driven H pump.
The light sensitive retinal -group covalently
bonded to a Lys-216 and block the channel.

Retinal can absorb a quantum of light (1) which
causes it to structure change (isomerization)
(2), releasing a proton into the external half of
the channel. Then rapidly isomerizes back,
picking up a proton from the inside half of the
channel (3).

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Result The absorption of light induces a series
of structural changes in the protein that cause
Step 1. a proton to move from the retinal group,
through a channel in the protein, to the cell
exterior. Step 2. the proton is replaced by
another proton transferred to the proteins from
the cytoplasm. Thus translocation of H generate
a H gradient across the plasma membrane. This
gradient is used by the ATP synthesizing enzyme
to make ATP.
Question
What is ATP, what is the function of ATP?
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Lys 216, with the retinal acts as
the gate in the channel.
Open-close gate is cis ? trans isomerisation
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Information exchange in the cell The cell lead
independent lives but they can communicate and
influence one another. In the most cases a
signaling network is based by Cell-surface
receptors - the membrane proteins The receptors
specifically bind the signaling molecule and then
initiate a response in the cell.
association
RECEPTOR LIGAND
? RECEPTOR-LIGAND
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Hormone Receptor is a membrane spanning protein
that recognizes and binds with specific hormone.
(?) Hormones - substances e.g. a peptide) are
chemical messengers in the body.
They
are produced by one tissue and conveyed by the
bloodstream to another

to effect physiological
activity, such as growth or metabolism. Only
those cells (target cells) containing a receptor
protein specific for the hormone can respond to
the hormone. Interaction between hormone and
receptor forms the
hormone receptor complex.
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Hormone Receptor Extracellular domains Some of
the residues exposed to the outside of the cell
interact with and bind the hormone.
Several distinctive variations in receptor
structure Some
receptors are simple, single-pass proteins

Others, such as the receptor for insulin, have
more than one subunit. Another class, which
includes the beta-adrenergic receptor, is
threaded through the membrane seven times.
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First Messengers Hormones mediate hormone action
within the cell. Hormones bind to receptors on
the surface of the cell, the message they convey
must be transferred into the cell by a variety of
mechanisms. Hormones are considered the first
messenger. Receptor molecules are neither
isolated by themselves nor fixed in one location
of the plasma membrane. Other integral membrane
proteins can interact with the receptor to
modulate its activity.

Some types of receptors cluster
together in the membrane after binding hormone.
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Result interaction of the hormone-bound
receptor with other membrane or cytoplasmic
proteins is the key to generation of second
messengers and transduction of the hormonal
signal. In all cases, the seemingly small signal
generated by hormone binding its receptor is
amplified within the cell into a cascade of
actions that changes the cell's physiologic
state.
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The second messenger - Cyclic AMP (cAMP)
is a molecule that is important in many
biological processes cAMP use for transferring
the effects of hormones which cannot get through
the cell membrane. (So hormone membrane
receptor complex works)
Its main purpose is the activation of protein
kinases (?)
it is also used to regulate the passage of Ca2
through ion channels.
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A protein kinase is an enzyme that modifies other
proteins by chemically adding phosphate groups to
them This usually results in a functional
change of the target protein by changing enzyme
activity, (decrease or increase in rate of a
chemical reaction),
cellular
location, or association with other proteins. Up
to 30 of all proteins may be modified by kinase
activity The human genome contains about 500
protein kinase genes they constitute about 2 of
all genes.
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How Cyclic AMP Second Messenger works The
intracellular concentration of cAMP is increased
or decreased by a variety of hormones The most
important effect of varibable concentrations of
cAMP is activation of a cAMP-dependent protein
kinase called protein kinase
A. Protein kinase A is nominally in an
catalytically-inactive state, but becomes active
when it binds cAMP. Upon activation, protein
kinase A phosphorylates a number of other
proteins, many of which are themselves enzymes
that are either activated or suppressed by being
phosphorylated. Such changes in enzymatic
activity within the cell clearly alter its state.
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Protein kinase A is comprised of two catalytic
and two regulatory subunits. In the absence of
cAMP, the regulatory subunits are bound to the
catalytic subunits and keep them inactive.
cAMP binds to
the regulatory subunits, ? it causes the
dissociation of active catalytic subunits from
the complex. ? provides signal amplification by
phosphorylation of a variety of proteins.
Mechanism of activation of protein kinase A by
cAMP.
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Tyrosine Kinase Second Messenger Systems The
receptors for several protein hormones are
themselves protein kinases which are switched on
by binding of hormone. The kinase activity
associated with such receptors results in
phosphorylation of tyrosine residues on other
proteins. Insulin is an example of a hormone
whose receptor is a tyrosine kinase.
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Tyrosine Kinase Second Messenger Systems The
receptors for several protein hormones are
themselves protein kinases which are switched on
by binding of hormone. The hormone binds to the
part of a receptor exposed on the cell's surface
and activates kinase domains located in the
cytoplasmic regions of the receptor. Thus the
receptor phosphorylates itself is the protein
kinase. Result The activated receptor
phosphorylates a variety of enzymes that become
activated or are inactivated upon
phosphorylation. Insulin is an example of a
hormone whose receptor is a tyrosine kinase.
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Fate of the Hormone-Receptor Complex Normal cell
function depends upon second messenger cascades
being transient events.
Indeed, a number of cancers are
associated with receptors that continually
stimulate second messenger systems.
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Ions play an essential role in cells They are
needed for signaling, catalysis of chemical
reactions, for maintaining the biologically
active structures of biopolymers. To enter
cells ions must permeate lipid membranes. This
process requires large activation energy
associated with transferring ions from the polar
aqueous environment to the nonpolar interior of
the membrane. What is the probability of a ion to
cross the membrane? The calculation showed that
it takes 1013 sec (about ten thousand year) for
one ion to pass through a purely lipid membrane.
Thus, a purely lipid membrane appears to be
practically impermeable for ions
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Porin - Transmembrane protein
ion transport is aided by ion channels, carriers
and pumps located in membranes.
Porin structure is highly regular wide close
cylinder from ß strands
close cylinder no free H-bond
donors and acceptors Cylinder comprises 16 very
long ß strands, diameter of a pore in its center
is 15Å
The side groups of polar residues face the pore.
Porin is responsible for transport of polar
molecules, but its selectivity is not too high
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Membrane water filled Channels
can be
simple pores such as "Porin"
If radius of the water channel (R 3Å) time to
pass the membrane about of a millisecond..
The channel sites can attract the ion.

For example, the presence of a positive charge
near the channel accelerates transport of
negatively charged ions and opposite
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Transmembrane photosynthetic reaction center
What is Photosynthesis?

is the process by which plants and some
bacteria use the energy from sunlight to produce
sugar,which later converts into ATP, the "fuel"
used by all living things.
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Light

6H2O 6CO2 ----------gt C6H12O6 6O2
Photosynthesis represents for the biosphere the
major source of ? Free energy (1017 kcal/year
stored) ? Carbon (1010 tons/year assimilated) ?
Oxygen
water and CO2 enter the cells of the leaf, and
the products of photosynthesis, sugar and oxygen,
leave the leaf.
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The process of energy conversion begins when
chlorophyll molecule is excited by a quantum of
light (a photon). It starts at the antenna
complex cluster of several hundred chlorophyll
molecules in the thylakoid membrane
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The antenna complex collected light energy and
directs it to the photochemical reaction
center.
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Photosynthetic reaction integral membrane centre
contains pigments such as chlorophyll, that
absorb light and transfer that light energy to
light-driven electron transfer reactions.

Chlorophyll gives leaves their green colour
The electron flow produced by the reaction center
chlorohylls is used to shuttle H ions across the
thylacoid membrane, setting up a potential mainly
used to produce ATP molecules.
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Crystal Structure Of Photosystem I A
Photosynthetic Reaction Center and Core Antenna
System From Cyanobacteria (Nobel prize 1988)
At the heart of each complex is an arrangement of
two sets of five transmembrane
a-helices, with quinone cofactors.
These cofactors catalyse the
photochemical transmembrane electron transfer
reaction that is the key to the photosynthetic
process.
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Home assignment The membrane proteins and
diseases (PowerPoint presentation)