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1. Details of Module and its Structure
Module Detail
Subject Name Botany
Paper Name Cell Biology
Module Name/Title Cell membrane and Cellular transport III
Module Id
Pre-requisites
Objectives To understand principle and mechanism of primary active transport Keywords Active transport, Electochemical gradient Primary active transport, Facilitated Diffusion, Carrier proteins for primary active
Structure of Module / Syllabus of a module (Define Topic / Sub-topic of module )
Passive transport
Passive transport < Introduction > < Active transport>, < Electochemical gradient> < primary active transport> < Facilitated Diffusion > < carrier proteins for primary active transport >
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2. 2. Development Team
Role Name Affiliation
National Coordinator
Subject Coordinator Dr Sujata Bhargava
Paper Coordinator Dr Nutan Malpathak
Content Writer/Author (CW) Dr Pradnya Kedari
Content Reviewer (CR)
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TABLE OF CONTENTS (for textual content)
1 Introduction
2 Active transport
4 Primary active transport
5 Carrier Proteins or pumps for Active Transport
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Cell membrane and cellular transport III
1. Introduction
Active transport is a transport of molecules against its concentration gradient. Molecules are carried from region of lower concentration to region of higher concentration. This process face resistance and so it has to utilise energy to overcome this resistance and carryout this transport. Therefore it is called as active transport. The process is also known as uphill transport. This energy is usually in the form of adenosine triphosphate (ATP). This ATP is utilized directly/ indirectly.
Cell need to move molecule - against concentration gradient too.
Active transport is a transport of a molecule which requires expenditure of energy to move solute against conc. gradient, using energy.
Active transport mechanisms use cell’s energy, usually in the form of Adenosine triphosphate (ATP).
In case of primary active transport ATP is utilized directly.
It is an uphill transport.
Let’s revise the difference between passive transport and active transport?
Active and passive transport are terms that are utilized in terms of plant, animals as well as human body.
2 Active transport
Active transport involves carrying a molecule against the concentration gradient, and it utilizes energy, whereas passive transport occurs according to concentration gradient and therefore it does not require any energy.
During active transport, as resistance is more it is called as uphill process, whereas passive transport face no resistance and therefore is referred as downhill process.
Some active transport mechanisms move small-molecular weight materials, such as ions, through the membrane. Other mechanisms transport much larger molecules.
An example of active transport is - uptake of salt by cells through a sodium pump
3 Electrochemical Gradient?
living systems, concentration gradients are more complex. In a cell, ions are constantly moving in and out of cells. They posses either positive or negative cgarge. Cell also contains proteins which do not move across the membrane. These proteins are
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mostly negatively charged. This creates an electrical gradient i.e difference of charge, across the plasma membrane. Interior of a living cell is electrically negative W. R. t. extracellular fluid.
At the same time, cells possess higher concentrations of potassium (K+) and lower concentrations of sodium (Na+) than the extracellular fluid. Therefore in a living cell, the concentration gradient of Na+ tends to drive it into the cell, and the electrical gradient of Na+ (a positive ion) also tends to drive it inward to the negatively charged interior. This makes the situation more complex for other elements such as potassium. As a living cell is electrically negative, electrical gradient of K+, a positive ion, also tends to drive it into the cell, but concentration of K+ ions is greater into the cell as compare to extracellular fluids. Therefore the concentration gradient of K+ tends to drive K+ out of the cell. Such combined gradient of concentration and electrical charge affects an ion and is called its electrochemical gradient.
If it’s a Fact that Potassium solution Injection is lethal. My question is - Why do you think a potassium solution injection is lethal? Answer to this question is that + + • Cell- have high K conc. In the cytoplasm, and it has high conc. of Na outside,
+ • Due to K Injection,thete is high conce of K+ ions outside the cell which cahnges electrochemical gradient. • Sodium/potassium potential from heart muscle, play important role in transmitting muscle contraction signal. • Due to change in this electrochemical gradient, heart fails to send the beating signal, and therefore person dies. • That is the reason why a potassium solution injection is lethal, but during heart surgeries, This K+ injections are used during in appropriate amount to stop the heart from beating during surgery.
Moving Against a Gradient
For a cell, it needs to spend an energy to move a substances against a concentration or electrochemical gradient. This energy is gained from hydrolysis of ATP which is generated through the cell’s metabolism.
Small substances are constantly passing through plasma membranes. It keeps on fluctuating the electrochemical gradient of a cell. There is special active transport mechanism followed by a cell to carry outs active transport process, which is known as pumps. Pumps work against this electrochemical gradient. Active transport maintains concentrations of ions and other substances needed by living cells in response to these passive movements. Cell needs to spend much of its supply of metabolic energy to maintain these processes.
(Most of a red blood cell’s metabolic energy is used to maintain the imbalance between exterior and interior sodium and potassium levels required by the cell.)
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Because active transport mechanisms depend on a cell’s metabolism for energy, they are sensitive to many metabolic poisons that interfere with the supply of ATP.
Primary active transport
There exist two mechanisms for the transport of small-molecular weight material and small molecules.
1. Primary active transport which moves ions across a membrane and creates a difference in charge across that membrane. This is directly dependent on ATP. 2. Secondary active transport which describes the movement of material that is due to the electrochemical gradient established by primary active transport that does not directly require ATP.
The cell makes use of membrane pumps to accomplish active transport.
Pumps can convert free energy into different forms, depending on which form is required by the cell at a given time.
This property makes membrane pumps a convenient choice for mediating active transport as they can provide the energy needed to initiate the transport.
The two main types of pumps employed by the cell are P-type ATPases and ATP-binding cassette transporters (eg-ABCs).
Both of these pumps are powered by ATP.
One method by which these pumps can perform active transport is by binding to ATP. This binding, followed by hydrolysis which induces a conformational change in the pump that allows bound ions to be transported across the cell membrane.
These pumps can also use active transport to establish favorable concentration gradients for separate transport processes.
For example, one pump can create a given concentration gradient by performing active transport on a certain ion, and then another pump can make use of this new concentration gradient and facilitat ion diffusion down the concentration gradient.
Thus the cell can couple active transport with passive transport in order to efficiently use the results of one process to drive another process to completion.
4 Carrier Proteins for Active Transport Membrane show certain mechanisms to adapt the process of transport against the concentration gradient. An important membrane adaption for active transport is the presence of specific carrier proteins or pumps. They facilitate this movement.
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There are three types of these proteins or transporters present on a membrane. Uniporter which carries one specific ion or molecule at a time. Symporter which carries two different ions or molecules at a time. Both the molecules move in the same direction. And Antiporter, which also carries two different ions or molecules ata time, but in different directions. All of these transporters can also transport small, uncharged organic molecules like glucose. These three types of carrier proteins are also found in facilitated diffusion, but they do not require ATP to work in that process. Some examples of pumps for active transport are Na+–K+ ATPase, which carries sodium and potassium ions, and H+–K+ ATPase, which carries hydrogen and potassium ions. Both of these are antiporter carrier proteins. Two other carrier proteins are Ca2+ ATPase and H+ ATPase, which carry only calcium and only hydrogen ions, respectively. Both are pumps. Sodium-potassium pump (Na+-K+ ATPase) are
The Na+-K+ pump helps it maintain this concentration by generating the necessary ion gradient.
Antiporter carrier proteins As Various cellular processes require K+, animal cell possess high amount of K + than Na+. An enzyme Na+-K+ ATPase are used to control this ion gradients in intracellular media They Actively transport most common ions i. e Na+ out and K+ into the cell • They Maintains the electrochemical gradient, Thus Most important pump in animals cells • It transfers 3 Na +:2 K+ at a time • Na+-K+ ATPase - exist in 2 forms, • These Forms depend on- Orientation, And Affinity for either Na+-K+ ions
Na+-K+ pump
The Na+-K+ pump, also known as the Na+-K+ ATPase, is an enzyme used by the cell to control the ion gradients in its intracellular media. As the name of the enzyme suggests, the ions it deals with are those of potassium and sodium, which are two of the more common ions present in living
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systems. In this case, the Na+-K+ pump hydrolyzes ATP in order to supply the energy needed to actively transport Na+ out of the cell while bringing K+ into the cell. It is for this reason that most animal cells tend to have a significantly higher concentration of potassium ions than sodium ions; the cell requires this concentration in order to be able to facilitate various cellular processes.
Mechanism of Na+-K+ ATPase is as follows
With the enzyme oriented towards the interior of the cell, the carrier has a high affinity for sodium ions. Three 3 Na+ ions bind to the protein. As a result of ATP is hydrolysis and low-energy phosphate group attaches to the carrier. This attachment results in a Change of conformation of these proteins, due to which the protein reorients. This re-orientation turns the carrier towards exterior side This Shape change results in the decreased affinity of protein for 3 Na+ and Increases the + affinity for 2 K , + Thus, 3 Na+ ions leave the carrier at the same time, 2 K ions attach to this carrier Now, Low-energy phosphate group detaches from the carrier, again resulting in conformation change which repositions the carrier to interior. + This New conformation, results in Decreased affinity for 2 K and Increased affinity for 3 Na+, + Thus, 2 K are released into the cytoplasm, and Carrier binds with 3 Na+ due to increased affinity to Na+. And so on…. For every three ions of sodium that move out, two ions of potassium move in. This results in the interior being slightly more negative relative to the exterior. This difference in charge is important in creating the conditions necessary for the secondary process. The sodium-potassium pump is, therefore, an electrogenic pump (i.e a pump that creates a charge imbalance), creating an electrical imbalance across the membrane and contributing to the membrane potential.
Two other such enzyme "pumps" that are homologs of the Na+-K+ pump are the Ca2+ ATPase and the H+-K+ ATPase. Ca2+ ATPase
Ca2+ ATPase ensures Transport of calcium ions (Ca2+) from normal cells to sarcoplasmic reticulum of muscle cells.
Calcium ions are needed by the muscle cells in order to function at peak efficiency; thus this pump ensures that they receive enough calcium ions to fulfill this requirement.
• These are An integral membrane protein • They are Comprised of a single polypeptide chain of 994 amino acid residues
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• These are ATP-powered Ca 2+ -pump • It’s a pump that transports only calcium • Ca + needed by muscles to function at peak efficiency
Lets c the Mechanism
These Plays role in muscle contraction Ca2+ is released from sarcoplasmic reticulum (SR) into muscle cells via Ca2+-release channel. Ca2+-ATPase, pumps back the released Ca2+ into the SR to cause relaxation. This pump can run when ATP and Ca2+ are available in the cytoplasm.
H+-K+ ATPase
H+-K+ ATPase pumps are Antiporter carrier proteins. H+-K+ ATPase are large quantities of protons (H+) into the stomach's gastric juices in order to keep the pH less than 1.0. Such a low pH is required because the stomach is responsible for digesting all the foods and fluids that enter it; thus its gastric juices need to be acidic enough to dissolve and break down anything that needs digesting.
These three pumps are all part of a family known as the P-type ATPases. They are called P-type ATPases because they all form a phosphorylated intermediate during their reactions with ATP. Hundreds upon hundreds of known homologs of these pumps exist within the P-type ATPases, and each plays a definite role in maintaining the functions of the cell.
Multidrug Resistant (MDR) Pumps
MDR Pumps are activated by ATP and are found in microorganisms such as bacteria and cancer cells.
The multidrug-resistance (MDR) pumps consist of large proteins that weave through the cell- surface membranes. They work to effectively monitor and prevent unwanted chemicals from entering the cell. Therefore, the microbes have ability to self-defend with MDR pumps. MDR pumps also prevent antibiotics from entering bacteria and chemotherapeutic agents from entering cancer cells. In addition, they are used to spit out the ones that might endanger the bacteria. Administering antibiotics with MDR inhibitors may be a way to circumvent the MDR pumps in bacteria. This explains the uncanny ability of bacteria to defend themselves against antibiotics. MDR pumps can be found in humans, where they have variety of roles that help drugs get to places they need to go. They are also important in membranes in the brain, digestive tract, liver, and kidneys because they move hormones into and out of cells through the membrane (Medicines by Design 15).
For instance, AcrB is a effective MDR in Gram-negative bacteria. It functions through its three asymmetric protomers, each of which exists as a particular conformation that corresponds to its function in the pump. The three stages are Access, Binding, and Extrusion. In the Access stage, substrates enter the protomer's vestibule, with the binding pocket still left in tact. In the Binding
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stage, the substrate remains in the vestibule, but the binding pocket expands to better hold the substrate. Lastly, in the Extrusion stage, the substrate exits via the removal of the central helix. AcrB serves to export a multitude of drugs out of the membrane, some of which include antiseptics, toxic compounds, and antibiotics. Vesicles
Vesicles are simply a bubble of fluid within a cell. Specifically, vesicles are a membrane bound sac that aid in transportation and storage of cellular waste or cellular products, metabolism, and buoyancy control. The membrane surrounding the vesicles shares many characteristics with the plasma membrane of the cell. As a result, the vesicle can fuse with the membrane in order to dispose of its contents.
Transport Vesicles
These are just a specific type of vesicle with a function of transport. They move molecules, products etc. between the Rough Endoplasmic Reticulum and the Golgi apparatus. Two types of proteins are synthesized on the ribosomes present on the Rough ER. From here the transport vesicles take the new proteins to the Golgi apparatus. At the Golgi apparatus the proteins age and mature in order to be transported to their final destination. The proteins always travel around the inside of the cell with transport vesicles.
Lipid Vesicles
Also known as liposomes, these vesicles are aqueous compartments that are surrounded by a lipid bilayer. These vesicles are used to monitor membrane permeability and to deliver chemicals to the cell. Lipid vesicles aid in determining the level of impermeability of a membrane to ions and polar molecules. Ions and polar molecules are trapped in the aqueous compartment of the liposome. The rate of the flow from the inner compartment of the vesicle to the outside solution during membrane transport determines the membrane's impermeability to the ions or polar molecules contained in the inner aqueous compartment. Liposomes are formed by sonicating phospholipids in the presence of an ion, polar molecules or a water soluble substance.
Coated Vesicles:
These are vesicles that form when emerging buds are detached from the cell membrane. Depending on the formation of the vesicle, the formation of the outer coat may form the vesicle, where in other cases, other enzymes are necessary to form the vesicle. It is an intracellular structure. The outer surface of the vesicle is covered in a lattice or cage like coating of the protein Clathrin.
Clathrin is a protein that forms a lattice shaped covering on the cytosolic side of the cellular membrane. These cytosolic sides are referred to as coated pits, the first stage of forming coated vesicles. It is made through a process of making subunits called triskelioins. The Triskelioins are a three pronged molecule with three N-terminus regions. They are known as heavy chains, about
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192kDa in weight and are bound to light chains, about 30kDa in weight. The invagination of the pit is during the first states of endocytosis and results in a clathrin coated vesicle. Clathrin can self assemble spontaneously and it plays a major role in deforming the budding vesicle. While clathrin is a major player in the deformation, there are other accessory proteins that aid in the assembly and dissembly of the coats.
COP I is another protein that coats vesicles during protein transportation. However, this vesicle coat transports from the Golgi apparatus to the Rough Endoplasm Reticulum. Since this is in the reverse of direction of normal transportation, it is known as a retrograde transport. The protein coat is made of large protein sub-complexes that possess 7 unique subunits.
The protein is known as an ADP Ribosylation Factor dependent adaptor protein. This basically means that they are regulators of vesicle biogenesis. They also aid in the selection of the proteins that are used for the carriers.
COP II is very similar to COP I in the fact that it transports proteins, however COP II transports from the Rough ER to the Golgi apparatus and is known anterograde transport. COP II has a coat that is made of 4 unique protein subunits. COP II has three different binding sites that it can complex with other proteins.
Both COP I and COP II are very active in membrane trafficking. They perform the necessary tasks of selecting the correct cargo for the proteins and they change the shape of the phospholipid bilayer into the correct buds and vesicles.
The anterograde and retrograde transport off set each other to help keep the flow of the membranes and secretions in balance. There is constant recycling going on to maintain the equal amounts of resident proteins in the different pathways. ABC Transporters
ABC transporters are a family of transport proteins that depend upon ATP binding for transport. ABC stands for ATP-Binding Cassette. ABC proteins have a particular molecular structure that includes two nucleotide binding domains where ATP binds. An example is the multidrug resistance protein (MDR1). This protein uses the energy of ATP hydrolysis to pump a wide variety of nonpolar drugs and toxins out of cells. It is so named because over-expression of this protein in tumor cells confers resistance to chemotherapy drugs.
A physiologically important member of the ABC transporter family is the protein CFTR. CFTR forms a Cl- channel that is expressed on the apical plasma membrane of many epithelial cells. CFTR is the protein that is defective in the genetic disorder cystic fibrosis. Unlike most ABC transporter proteins that use the energy of ATP hydrolysis to pump substances across the membrane and out of cells, CFTR works as an ion channel that is regulated by both phosphorylation and ATP binding.
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CFTR plays a key role in the secretion of ions and water across epithelia (see page on Epithelial Transport). Some bacterial toxins cause unregulated activity of CFTR, resulting in excessive secretion in the small intestine which causes diarrhea. In cystic fibrosis, CFTR channels are defective or absent, leading to decreased secretion, which causes pathology in the lungs and digestive system.
So this was about primary Active transport which describes the process of transport of specific substances. This transport is directly coupled to ATP hydrolysis. Because the energy for transport is derived from ATP hydrolysis, these transporters effectively move substances in one direction, regardless of the concentration gradient.
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