Bio test 3 - Cell Mechanics

Plasma Membrane

Also known as the cell membrane

All cells and organelles are surrounded by a flexible membrane

Organs

specialized structures in the body that perform specific life processes

Organelles

specialized structures inside the cell that perform specific cellular processes

often surrounded by a membrane

Cell Fractionation

A method of separating cell parts to study their function

Homogenization

part of cell fractionation

disruption of cell membrane without damaging organelle

breaks the cell apart but doesn’t identify what is inside (1st step)

Centrifuge

instrument that spins at high speeds to separate contents by density

List and explain the 4 methods of homogenization

1. break cell apart with **high frequency sound**
2. use **mild detergent** to make holes in plasma membrane
3. force cells through small hole using **high pressure**
4. **shear** cells between a close-fitting rotating plunger and the thick walls of a glass vessel

how does mild detergent work for homogenization

both phospholipid membrane and detergent have hydrophobi and hydrophillic properties which creates holes in membrane and membrane disintegrates

what is the result of homogenization called

homogenate or an extract

describe the homogenate

thick soup, contains large and small molecules from the cytosol, such as enzymes, ribosomes, and metabolites, as well as all the membrane bound organelles

does homogenization leave the membrane bound organelles intact

yes

what are the 2 substances in centrifugation

supernatant and pellet

What does the pellet contain

larger, more dense components

what does the supernatant contain

lighter, suspended in liquid above the pellet

Steps to Cell Fractionation

1. Homogenize
2. Centrifuge

* Pellet: larger, more dense components
* Supernatant: lighter, suspended in liquid above the pellet


3. Decant supernatant (the act of pouring, syphoning, or draining the top or bottom portion of a liquid)
4. Repeat centrifugation at higher speeds to separate into smaller components

Which centrifuged layer (supernatant or pellet) is transferred to the next fractionation step? Why?

supernatant is transferred to get smaller components of the cell in the next centrifugation

Which fractionation step produces the least dense organelles?

the last centrifugation

What is the difference between each of the centrifugation steps?

Why is this difference needed?

time and g force are different, they increases with every set of spins to get smaller components

What is in the pellet after each centrifugation

1. nuclei and cellular debris
2. mitochondria (and chloroplasts for plants)
3. microsomes (pieces of plasma membrane and cell’s internal membrane)
4. ribosomes

What is the Fluid Mosaic Model used to help understand

to understand membrane structure

Who developed the fluid mosaic model

Model developed by Singer and Nicolson (1972)

what does the ‘fluid’ part describe

Fluid implies movement on membrane

what does the ‘mosaic’ part describe

Mosaic implies that the membrane consists of many different molecules

Membrane Composition

Mosaic

Plasma membrane is composed of many different molecules:

* Phospholipids
* Membrane Proteins

- Integral

- Peripheral

* Carbohydrates
* Cholesterol

Plasma Membrane Structure

Cell membrane made of phospholipid

Phospholipids also form the membrane around organelles

Phospholipid Bilayer

Bilayer = 2 layers

composed of phospholipids

what is each layer called

Each layer is called a leaflet

amphipathic

hydrophobic and hydrophilic properties

Where is water in the phospholipid bilayer?

Water is on the intracellular and extracellular side

* Cell is in a water (polar) environment
* Cytoplasm (cell interior) is also a water environment

How are the phospholipids arranged

Phospholipid arranged so that hydrophobic tails do not face water

* Hydrophobic tails can not face outside or inside the cell
* Hydrophobic tails face inwards forming a hydrophobic core
* Hydrophilic heads face outwards

Two Types of Membrane Proteins

Integral and Peripheral

Integral Proteins Classification/Structural classes of integral membrane proteins

Polytopic Transmembrane

• Single-pass

• Multi-pass

Monotopic

What are the two types of polytopic transmembrane integral proteins

single-pass and multi-pass

Polytopic

faces both sides of membrane

Transmembrane

spans entire phospholipid bilayer

Single-pass

crosses membrane once

Multi-pass

crosses membrane several times

Monotopic

associated with membrane on one side (e.g. one leaflet), does not span entire bilayer

Integral Proteins Function

* Receptor
* Recognition
* Transport

• Channel

• Carrier / Pump

* Cell adhesion

• Anchoring

• Occluding

• Channel forming

Receptor Protein

Has a binding site for the ligand

Recognition Protein

Glycoprotein: surface carbohydrate groups help identify cell (e.g. antigens)

Two types of transport proteins

Channel and Carrier/Pump

Channel Proteins

* Act like tunnels
* Molecules move through protein passively (no energy involved)
* Moves small molecules or charged ions

Two types of Channel Proteins

Un-gated (leak channels): always opened

Gated (open or closed position): have open and closed conformations - Changes stimulated by changes in external environme

Carrier Proteins / Pumps

Acts like a turnstile or revolving door

Undergo conformational change to allow molecules through

Difference between carrier and pump proteins

Carrier proteins are typically molecules that bind to other compounds so as to facilitate passage through a membrane. On the other hand, a pump is a protein channel that relies on a gradient (usually chemiosmotic eg electrolytes) for action. Both turnstiles

What is cell adhesion also known as

cell junctions

Cell Adhesion

Structures that connect cell to cell

Allow cells to adhere to each other

Types of cell adhesion

Channel-forming junction

Occluding junction

Anchoring junction

What are channel forming junctions known as

gap junctions

Gap junctions

create gaps that connect animal cells, membrane proteins from adjacent cells line up to create a channel, like 2 channel proteins

What are occluding junctions also known as

Tight Junctions

Tight junctions

Forms impermeable barrier between cells, membrane proteins create tight junctions, act like staples so that substances don’t leak in between cells

Where in your body, would tight junctions be crucial?

blood-brain barrier, gastrointestinal tract (need transport proteins to absorb food, it can’t go through cells, prevents pathogens from going through)

What are anchoring junctions also known as

Desmosome (a type of anchoring junction)

Desmosome

Desmosomes bind to desmosomes on adjacent cells

Attached to cytoskeleton

Helps resist shearing force

flexible and can move to precent ripping of cells

Peripheral Membrane Proteins

Bound non-covalently to either surface of the membrane

Peripheral Proteins Classification

* Extracellular
* Intracellular

Peripheral Proteins Function

* Communication
* Structural support

Which function goes with which classification?

Extracellular - communication

intracellular - structural support

Extracellular Peripheral Protein

Communication

* Located on outer leaflet and surface
* Receptor and recognition proteins (which can also be integral)

Extracellular Matrix (ECM): Components

* A matrix of glycoproteins
* Secreted by cells
* Varies with type of tissue

Extracellular Matrix (ECM): Function

Supports cell structure

Anchors cell

Separates tissues

Functions in cell signalling

Intracellular Peripheral Protein

Structural

* Cytoskeletal protein
* Located on inner membrane surface
* Attached to cytoskeleton of cell
* Immobilized (anchored) on membrane

Cytoskeleton

A network of fibers extending throughout the cytoplasm

Dynamic: can be quickly dismantled and reassembled in a new location

Cytoskeleton Components

Microfilament: actin

Intermediate filament

Microtubules: tubulin

Microfilament structure

2 intertwined strands of actin

intermediate filament structure

Fibrous protein supercoiled

microtubule structure

Hollow tube of 13 tubulin columns

Based on the picture, where is the nuclei and where is the cytoskeleton

Nuclei (green)

Cytoskeleton: microfilaments (actin) (purple), microtubules (yellow)

what are the two objects that carbohydrates attach themselves to

proteins and lipid

where are carbohydrates found in the membrane

Glycoproteins, glycolipids

Found on Extracellular side

glycoprotein

carbohydrate + protein

glycolipid

carbohydrate + lipid (phospholipid)

What is the function of cell surface carbohydrates?

* identifies the cell (like a name) helping other cells recognize it
* acts as a signal for communication

Of the component of the plasma membrane that was just studied, which would have an affect on the fluidity of the membrane?

* Phospholipids
* Proteins
* Carbohydrates

phospholipids - depending on saturation changes fluidity

Membrane fluidity is affected by:

* Saturation of fatty acid
* Hydrophobic restrictions
* Cholesterol and temperature

Saturation of Fatty Acid

Double bonds bends fatty acid chains preventing phospholipids from tight packing

How can phospholipids move in the membrane (5 ways)

lateral diffusion, rotation, swing, flexion, transverse diffusion

Lateral Diffusion

movement across same leaflet (phospholipids transpose with neighboring molecules)

Rotation

is when an individual molecule rotates quickly around its axis

Swing

from side-to-side

Flexion

contraction movement

how do hydrophoblic restrictions affect fluidity

make the transverse diffusion rare because the polar heads have to pass through the hydrophobic tails

Transverse Diffusion

flip-flop

movement from one leaflet to the other

rare because because the hydrophilic head of phospholipid must go cross the hydrophobic core to get to the leaflet

what enzyme allows for transverse diffusion

facilitated by enzyme flippase

Properties of Cholesterol

Large molecular size, Nonpolar

How does the large molecule size work in the bilayer membrane

Can interrupt intermolecular forces of attraction

How does the non-polar property work in the bilayer membrane

Stabilizes hydrophobic interactions

Explain which property of cholesterol plays a more significant role at: Low temperature

large molecule, interrupts hydrophobic interaction causing the bilayer to be flexible

Explain which property of cholesterol plays a more significant role at: High temperature

non-polar, stabilizes hydrophobic interactions by attracting the phospholipids making the bilayer more rigid

Why is it advantageous for a cell to have cholesterol?

wider range of temperatures, stops membrane from falling apart in high temps and stops from freezing at low temps

Is the membrane symmetrical

no, there is asymmetry

How is there asymmetry

Each leaflet has a different composition

Leaflet facing the intracellular side has different components compared to the extracellular side

What does an asymmetrical membrane bilayer imply

it implies that the two sides of membrane are structurally and functionally different

How is the asymmetry maintained

Restrictions in the transverse (flip-flop) motion help to maintain this asymmetry

If you were given an illustration of a cross-section of a cell membrane, describe two things that would help you identify the side that faces the outside environment.

glycoproteins, glycolipids, fibers of the extracellular matrix

Basic Membrane Function

* Maintain internal cell environment at a steady state regardless of changes in the external environment
* Similar to the concept of homeostasis in the human body but this is at the cellular level
* Acts as a selective barrier regulating the movement of substances into and out of the cell

Solute vs. Solvent

solute is what dissolves in a solvent

Concentration

The amount of solute dissolved in the solvent, refers to solute