Cheng Xiaoli Zhengzhou University

Cheng Xiaoli Zhengzhou University

Chapter 3

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Content s 3.3.1 General

Introduction 3.3.2 Passive Transport 3.3.3 Facilitated Diffusion 3.3.4 Active Transport 3.3.5 Several topic for ion pumps Cheng Xiaoli Zhengzhou University

3.3.1 General Introduction

• Transport of nutrients, ions, and excretory substances from one side to the other is a major function of the cell membrane. A number of different means have been developed to fulfill this function. • The interior of the lipid bilayer is hydrophobic and blocks the passage of almost all water-soluble molecules. But cell live and grow by exchanging molecules with their environment, so that various water-soluble molecules must be able to cross the plasma membrane. A few of these solutes can simply diffuse across the lipid bilayer, but the vast Cheng Xiaoli Zhengzhou University

3.3.1 General Introduction

majority can not. Instead, their transfer depends on membrane transport proteins that span the membrane and provide private passages across it for specific substances. • Movements of molecules “downhill” from a region of high concentration to a region of low concentration occur spontaneously, provided a pathway exits. Such movements are called passive, because they need no other driving force. Cheng Xiaoli Zhengzhou University

3.3.1 General Introduction • To move a solute against its concentration

gradient, however, a transport protein has to do work: it has to drive the “uphill” flow by coupling it to some other process that provides energy. Transmembrane solute movement driven in this way is termed active transport. • Why transport ? • Ensure the essential molecular enter the cell • Remove waste • maintain a constant internal environment Cheng Xiaoli Zhengzhou University

3.3.1 General Introduction

The meaning of selective permeability • Plasma membrane is a selective permeable barrier between the cell and the extracellular environment. • its selective permeability ensures that essential molecules such as ions, glucose, amino acids, and lipids readily enter the cell, metabolic intermediates remain in the cell, and waste compounds leave the cell. • The selective permeability of plasma membrane allows the cell to maintain a constant internal environment. Cheng Xiaoli Zhengzhou University

3.3.2. Passive Transport • Simple diffusion. • The simplest form of transport is passive diffusion. It is a real freebie; it does not even need helpers. • Fat-soluble molecules and small uncharged molecules can pass directly through the lipid bilayer by simple diffusion. high concentration Concentration gradient

low concentration Cheng Xiaoli Zhengzhou University

3.3.2. Passive Transport • Simple diffusion Few molecules, such as Oxygen, carbon dioxide, nitrogen (O 2, CO 2, N 2 ), diffuse easily through membrane because they have no charge (partial or complete) to interact with water. such urea and ethanol, can diffuse across an artificial membranes composed of pure phospholipid or of phospholipid and cholesterol. hydrophilic hydrophobic molecule molecule • Hydrophobic molecules (oils) have also no trouble permeating

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3.3.2. Passive Transport • Simple diffusion In the process, no metabolic energy is expended, no specific transport proteins need, movement down its chemical concentration gradient, called passive diffusion, or simple diffusion. High Concentration

Small gaseous molecules

O2 CO2 N2

Low Concentration

permeable

Small glycerolpermeable uncharged polar ethanol molecules relative permeability of a pure phospholipid bilayer Cheng Xiaoli Zhengzhou University

3.3.2. Passive Transport • Simple diffusion • Passive diffusion is driven by the thermodynamic action. The diffusion rate of any substance is determined by its concentration gradient across the layer, its hydrophobicity, its size and any electric potential across the membrane affected charged molecules. The greater concentration gradient of the substance across the layer, or the higher its hydrophobicity, or the smaller molecule, the higher its diffusion rate is. Cheng Xiaoli Zhengzhou University

3.3.2. Passive Transport • Simple diffusion Small uncharged polar molecules Charged polar molecules Ions

Urea, Water

Slightly permeable

Amino acids ATP Nucleic acids protein H+ Na+ HCO3Mg2+

ClCa2+ K+

Impermeable

Impermeable

relative permeability of a pure phospholipid bilayer Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport • In most human cells there is an unequal distribution of ions: Na + 10 mM inside; 150 mM outside.

Na 150 mM

Cl Na 10 mM organic anions

- 70 mV K 5 mM

K + 150 mM inside; 5 mM •outside. anions: proteins inside; Cl outside

K 150 mM

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3.3.3. Facilitated Transport Distribution of ions across the cell membrane

Component K+ (potassium ion)

outside (mM) 5

Na+ (Sodium ion) Ca+2 (calcium ion)

150 1.8

Cl- (chlorine ion) Mg+2 (magnesium ion)

110 1.5

Amino acids, proteins

9

inside (mM) 150

10 ＜ 0.0005 3 0.8 138

• So the cell membrane is called selective permeability barrier that maintains distinct internal and external cellular environments. • It must get supports otherwise those large molecules and ions do not penetrate bilayer. Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport

Overview • The kinetics of facilitated (with a helper) transport is different from those of simple diffusion. In the latter, the rate of diffusion is proportional to the concentration of the diffusing molecules; the more of them, the more of them rate o f tran sp o rt will diffuse across the membrane per unit time. fa cilitated diffu s io n • The rate of facilitated diffusion is far higher than passive diffusion.

s im p le d iffus io n

Cheng Xiaoli [c on c. tranZhengzhou sp orted mUniversity olecule]

3.3.3. Facilitated Transport

Overview • The rate of facilitated diffusion is limited by the availability of the uniporter molecules rather than throughout the phospholipid bilayer. Once all the helpers are saturated, the increasing concentration of diffusing molecules will only increase a waiting line for the helper and will not increase rate of transport. • Consequently, there is a maximum transport rate Vmax that is achieved when the concentration gradient across the membrane is very large and each uniporter is working at its maximal rate. Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport Overview

• Facilitated transport is specific. Each uniporter transports only a single species of molecule or a single group of closely related molecules.

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3.3.3. Facilitated Transport • Ion channels The simplest form of a helper-facilitator is an ion channel. • Ion channels are usually small proteins that span the membrane and have central water-filled pores and its outside surface is hydrophobic and the inside hydrophilic. hydrophobic

hydrophilic

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3.3.3. Facilitated Transport • Ion channels •Most ion channels are gated and open transiently in response to a specific stimulus, such as a change in membrane potential or the binding of a ligand or a chemical (phosphorylation) or electrical stimulus . Ligand-Gated channels

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3.3.3. Facilitated Transport • Ion channels • Voltage-gated channels-– respond to voltage changes in membrane potential. ＋ ＋＋ ＋ ＋ ＋ ＋ ＋ ＋ － ＋

－ － － －

－ － － － － － －

＋ ＋

＋ －

－

－

Electrochemical gradient Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport • Ion channels • Ion channels have two distinct characters: high transport speed (high to 106 inos/s) and specific selectivity. The speed is above 1000 times higher than any carrier protein. • The forces driving the transport are due to concentration gradients and electric potential gradients. • Above 100 kinds of ion channels had be discovered on various cell’s plasma membrane. Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport • Transporters

(Carriers) • Transporters consist in almost all biomembrane and move a wide variety of ions and molecules across the membranes. • Don’t like channel protein, it is necessary that molecule or ion transported link to the transporter. Each kind of transporter links to special solution molecule, and introduces them from the side to another side of the membrane by the changing the conformation of itself. Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport

• Transporters (Carriers) • Three types transporters have been identified Uniporters Transported molecule transport a single type of molecule down its concentration gradient via • facilitated Facilitated diffusion requires no external diffusion. source of energy.

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3.3.3. Facilitated Transport

• Transporters (Carriers) Antiporters and symporters couple the movement of one type of ion or molecule against its concen-tration gradient with the movement of one or more different ions down its concentration gradient. transported molecule

Symport Cheng Xiaoli Zhengzhou University

3.3.3. Facilitated Transport

• Transporters (Carriers) For example , glucose, which is needed in large amounts by cells for energy, is one substance commonly cross the plasma membrane into cells with the aid of uniporters.

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3.3.3. Facilitated Transport

• Transporters (Carriers) Glucose transport also can catalyze the net export of glucose from the cytosol to the extracellular medium exterior when the glucose concentration is higher inside the cell than outside.

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3.3.3. Facilitated Transport

• Both carriers and channels are multipass tran-smembrane proteins. • A channel protein discriminates mainly on the basis of size and electric charge: if the channel is open, molecules small enough and carrying the appropriate charge can slip through, as though an open, but narrow, • A carrier proteintrapdoor. acts more like a turnstile: it allows passage only to solute molecules that fit into a binding site on the protein, and it transfers these molecules across the membrane one at a

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3.3.4. Active Transport

Often the transport has to happen in the direction opposite to the electrochemical or concentration gradient. In order to accom-plish this, membranes have evolved elaborate schemes to pump the substance ( ions tration to a compartment with or moleciles) from the area of higher smaller concen-tration. All these schemes concencost the cell energy (hydrolysis of ATP or light energy) and thus are called active transport. It is sensitive to inhibi-tors that stop ATP synthesis. Cheng Xiaoli Zhengzhou University

3.3.4. Active Transport

ATP-powered pumps

• ATP-powered pump is an ATPase that use the energy of ATP hydrolysis to move ions or small molecules across a membrane against a chemical concentration or electric potential gradient or both.

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3.3.4. Active Transport

ATP-powered pumps

• All ATP-powered pumps are transmembrane proteins with one or more binding sites for ATP located on the cytosolic face of the membrane. • Although these proteins commonly are called ATPase , they normally do not hydrolyze ATP into ADP and Pi unless ions or other molecules are simultaneously transported.

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3.3.4. Active Transport • There are four classes ATP-powered pumps: -- P- class ion pumps -- F-/ V- class ion pumps -- ABC (ATP-binding cassettes) superfamily

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3.3.4. Active Transport • The generally characteristics of ATPpowered pumps: -- are transmembrane proteins -have one or more ATP binding site (all in cytosolic side) -- coupling ATP hydrolysis/synthesis and ion transport

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3.3.4. Active Transport Four subunits of P-class pumps • H+ pump: plants, fungi, bacteria cell memb. • Na+ / K+ pump: animal cells membrane • H+ / K+ pump: mammalian stomach cells • Ca2+ pump: sarcoplasmic reticulum membrane in muscle cells

• during the transport process, at least one of the αsubunits is phosphorylated (hence the name Pclass pumps) and the transported ions are thought to move through the phosphorylated subunit. Cheng Xiaoli Zhengzhou University

3.3.4. Active Transport

P-class pumps functions • Maintains the low Na+ and high K+concentrations of animal cell Na 150 mM

Cl

• Pump Ca2+ into ER in muscle • Pump Ca out of the cytosol into the external medium 2+

Na 10 mM organic anions - 70 mV K 5 mM

K 150 mM

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3.3.4. Active Transport

F- / V- class proton pumps • F- and V- class pumps are similar to one another but unrelated to and more complicated than Pclass pumps. All known F- and V- class pumps transport only protons, in a process that does not involve a phosphoprotein intermediate. Na -K ATPase

• V- class pumps generally function to maintain the low pH of lysosomal and other acidic vesicles in animal H cells by pumping protons from the cytosolic to the exoplasm against a proton electrochemical gradient.

+

ATP H+ ADP+Pi

+

+

ATP Na+

K+

ADP+Pi

lysosome + + + H+

H + H+

H+ ATPase

ATP

H+

ADP+Pi

ATP Cheng Xiaoli Zhengzhou University

3.3.4. Active Transport

F-class proton pumps

• On Inner mitochondria membranes • On Bacterial plasma membrane • In contrast to V pumps, F pumps generally function to power the synthesis of ATP from ADP and Pi by movement of protons from the exoplasmic to the cytosolic face of the membrane down the proton electrochemical gradient.

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3.3.4. Active Transport

ABC superfamily

– Two transmembrane (T) domains: passage – Two cytosolic (A) domains: ATP-binding

ABC superfamily functions • Transport amino acid, sugar, peptide transporters (Bacterial ) • Transport phospholipids, cholesterol, lipophilic drugs, other small molecules Cheng Xiaoli Zhengzhou University

3.3.4. Active Transport

• Cotransporters

• Cotransporters mediate coupled reactions in which an energetically unfavorable reaction (uphill movement of molecules) is coupled to an energetically favorable reaction (symporter and antiporter). • Here, ATP pump use energy from hydrolysis of ATP, whereas cotransporters use the energy stored in an electrochemical gradient. This latter process sometimes is referred to as secondary active transport. Cheng Xiaoli Zhengzhou University

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Na+-driven glucose symport

Passive transport

Cheng Xiaoli Zhengzhou University Transport of glucose into and out of intestinal epithelial cells.

3.3.5. Several topic for ion pumps

The specific ionic composition of the cytosol usually differs greatly from that of the surrounding extracellular fluid. Invertebrate(Squid axon)

vertebrate (mammalian)

ion

Cell(mmol/L)

Blood (mmol/L)

K+

400

20

K+

139

4

Na+

50

440

Na+

12

145

Cl-

40-150

560

Cl-

4

116

Ca2+

0.0003

10

Ca2+

0.0002

1.8

Mg2+

0.8

1.5

X-

138

9

Mg2+ X- -

300-400

5-10

ion

Cell(mmol/L)

Blood (mmol/L)

X : represents proteins, which have a net negative charge at theCheng neutral Xiaoli pH Zhengzhou of blood and University cells.

3.3.5. Several topic for ion pumps

The ion pumps are largely responsible for establishing and maintaining the usual ionic gradients across the plasma and intracellular membranes. Cells expend considerable energy to carry out this task. For example, to transport ions, up to 25% of the ATP produced by nerve and kidney cells is consumed, and up to 50% of that in human erythrocytes; in both cases, most of this ATP is used to power the Na+-K+ pump. Cheng Xiaoli Zhengzhou University

3.3.5. Several topic for ion pumps

Studies on the effects of poisons Na 150 mM

Cl Na 10 mM organic anions - 70 mV

K 5 mM

Na 1 50 mM mM 75

K 150 mM

In cells treated with poisons K 5m M 75 mM

Cl Na 10 m M 75 mM organic anions

- 70 mV K 150 mM mM 75

75 mM

Eventually treated cells Die

Cells die: partly because protein synthesis requires a high concentration of K+ ions and partly because in the absence of a Na+ gradient across the cell membrane, a cell cannot import certain nutrients such as amino Cheng Xiaoli Zhengzhou University

3.3.5. Several topic for ion pumps

• Na / K - ATPase

Na/K ATPase is an important P-class ion pump to maintain the intracellular Na+ and K+ concentrations in animal cells.

• The Na/K-ATPase uses the energy of ATP hydrolysis to pump three Na+ ions out of the cell and two K+ ions into the cytosol against their electrochemical gradient. This enzyme is an important example of Cheng Xiaoli Zhengzhou University primary active transport.

3.3.5. Several topic for ion pumps

• Na/K-ATPase

Here the energy of a phosphate (shown in red) is used to exchange sodium atoms for potassium atoms. Cheng Xiaoli Zhengzhou University

3.3.5. Several topic for ion pumps Highaffinity binding sites

The mechanism of action of Na/KATPase

Sodium

Highaffinity binding sites

Potassium

lowaffinit y bindin g sites

lowaffinit y bindin g sites

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3.3.5. Several topic for ion pumps 2+

• Ca ATPase

•ATP- dependent Ca 2+ -pump •P-class ion pump

Ca2+

Ca2+

Ca2+ Ca2+ Ca2+Ca2+

• transport Ca2+ out of the cell or into intracellular Ca 2+ storage ag ainst its Ca2+ concentration gradi ent by using the chemical energy derived from ATP hydrolysis • help maintain the concentration of free Ca2+ in the cytosol at a low l evel

Ca2+

Ca 2+

ADP+Pi

ATP

Ca2+

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3.3.5. Several topic for ion pumps 2+

• Ca ATPase

Ca2+

Ca2+

• The catalytic cycle of ATPase can be reversed after a Ca2+ gr adient is formed across the ves icular membrane • During the reversal, Ca2+ leaves the vesicle through the ATPase and ATP is synthesized f rom ADP and Pi using the energ y derived from the Ca2+ concent ration gradient

Ca2+

Ca2+

Ca2+

Ca2+

Ca2+

ATP ADP+Pi

Ca2+

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3.3.5. Several topic for ion pumps 2+

• Ca ATPase

•Ca 2+ high concentration in the lumen of the ER serves as the major internal reservoir form which Ca 2+ is released to the cytosol. Skeletal and cardiac muscle contraction is activated when Ca 2+ is released from the SR lumen to the cytosol via 2+ Zhengzhou University theCheng CaXiaoli release

Key Points • Please explain the following terms: simple diffusion, facilitated diffusion, ion channels, Na/K-ATPase, symport, uniport, antiport and Cotransporter • Please compare the features distinguish uniport transport from passive difusion

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