BIOL 0500 Midterm #2

eukaryotic cells

• the smallest structural and functional unit of an organism capable of sustaining life

• eukaryotic cells have a distinct, membrane-bound nucleus that stores genetic information in the form of linear DNA

• work together with each other and other cells to form tissues

• have unique structures tailored to their specific functions

• come in different morphology and sizes

tissues

• groups of similarly functioning cells that work together to carry out a common function

• types: epithelial, muscular, connective, and neural

laser microscopes

• magnify species up to 1000 fold

• limited in their resolution (generally reveal structures up to 200nm)

• need an electron microscope to see structures with more detail

transmission electron microscope (TEM)

• uses a beam of electrons to scan very thin slices of tissue and isolated cells

• the electron gun sits at the base of the microscope, sending electrons shooting up in the following order: condenser lens>specimen>objective lens>projector lens>viewing screen

• staining tissues/cells with heavy metals provides contrast

• magnifies up to 1 million-fold and reveals structures as small as 1nm (200 times smaller than conventional)

plasma membrane

• the 5-10 nm thick outer boundary of the cell made up of 2 hydrophilic layers (leaflets) sandwiching a hydrophobic core

• higher ratio of cell membrane to cytoplasm to facilitate transport of materials in and out of the cell

• higher ratio of membrane surface area to cell volume means more efficient movement of resources, heat and waste

• embedded within the phospholipid bilayer are transmembrane proteins and other types of proteins and glycolipids

• the membrane also has several receptors that allow it to respond to signals from outside the cell

• membrane lipidd to protein ratio varies (e.g., myelin membrane is 80% lipid, while inner mitochondrial membrane is 75% protein)

plasma membrane roles

• “pc racisms”: protection, cellular communication, recognition of cells, attachment, cellular contents, identity, shape, movement of cells, selective barrier

• primarily allows for diffusion and active transport of materials

• the membrane is flexible yet selective due to its unique phospholipid composition and series of membrane proteins

membrane bound organelles

• GAMER LPN: golgi apparatus, mitochondria, endoplasmic reticulum, lysosomes, peroxisomes, nucleus

• organelles account for ~50% of the cell volume

• membranes allow organelles to carry out unique functions

• membranes are also reaction sites (e.g., ETC in mitochondria and phospholipid generation in endoplasmic reticulum)

nucleus structure

• function is to contain the cell’s genetic information in the form of chromatin and synthesize rRNAs

• is enclosed by a phospholipid bilayer that controls what goes in and out of the nucleus

• the nuclear membrane has pores through which molecules like mRNA pass through (DNA is too big)

• is physically attached to and continuous (contiguous) with the endoplasmic reticulum

• the nuclear envelope is comprised of nuclear protein complexes

• the nucleolus is a dark region within the nucleus responsible for rRNA synthesis

• comprises ~6% of total cell volume

endoplasmic reticulum

• flattened interconnected membrane tubules and sheets that extend from the nucleus into the cytoplasm, often continuous throughout the cell

• channels transport substances through the cell and provide a surface for chemical reactions to occur

• endoplasmic-reticulum derived vesicles fuse with the Golgi apparatus or plasma membrane

• the ER membrane synthesizes most lipids and proteins for distribution to organelles and plasma membrane

• likely gave rise to Golgi, peroxisomes, lysosomes, and endosomes, and supplies their lipids and proteins

• comprises ~12% of total cell volume

smooth endoplasmic reticulum

• lacks ribosomes, producing lipids, cholesterol, and fatty acids rather than proteins

• is uncommon in some cells but plentiful in some specialized cells

• in cells that produce steroid hormones (e.g., the adrenal gland), the smooth ER make cholesterol that is used to synthesize steroids

• is abundant in cells that detoxify molecules (e.g., in the liver where cytochrome 450 in the ER membrane oxidizes drugs)

• SER controls cellular calcium levels

sarcoplasmic reticulum

• the smooth endoplasmic reticulum of muscle cells

• Ca (2+) ions released from the SR lumen causes sarcomere contractions

• the SR lumen sequesters calcium ions, causing the relaxation of sarcomeres

rough endoplasmic reticulum

• the polyribosomes embedded in its surface synthesizes proteins of the ER, Golgi, endosome, lysosome, plasma membrane, and cell exterior

• soluble proteins enter the ER lumen and can stay there but usually travel to lumens of other organelles

• membrane proteins enter the ER membrane and can stay there but most travel to membranes of another compartment

• proteins used for extracellular purposes are sent to the Golgi apparatus

Golgi apparatus

• stacks of flattened, membrane-enclosed sacs found in eukaryotic cells

• Golgi apparatus membranes are asymmetrical with higher concentrations of lipids in the cytosolic leaflet since the Golgi bends toward the cytosol to form transport vesicles

• processes the molecular products of the cell (phospholipids, triglycerides, and proteins) before sorting them for transport to other sites (usually via vesicle transport)

• interacts with the ER and plasma membrane through vesicles

• redistributes lipids before they reach the plasma membrane

• pH decreases from the cis to trans Golgi network to facilitate enzyme driven protein modification, sorting, and packaging

• comprises ~3% of total cell volume

Golgi transport

• vesicles from the endoplasmic reticulum enter the cis Golgi, move t the medial Golgi, and exit at the trans Golgi

• Golgi compartments contain different enzymes and have different functions

• proteins are modified as they pass through the Golgi (e.g., oligosaccharides transferred from dolichol in ER are phosphorylated in cis Golgi)

Golgi compartments

• cis: the entry face of the Golgi apparatus found near the ER; acts as the receiving statin for the Golgi complex, receiving vesicles from the ER

• medial: located between the cis and trans faces, the region contains enzymes responsible for intermediate protein modifications (e.g., glycosylation, phosphorylation, etc.)

• trans: final, most acidic processing compartment that sorts and packages mature proteins and lipids into vesicles for transport to lysosomes, plasma membrane, etc.

• increased acidity as you move along helps sorting receptors bind cargo correctly, and helps regulate enzyme activity for glycosylation

• note that proteins for export maintain their orientation from the ER, through Golgi compartments and to the plasma membrane (i.e., the lumen side remains facing inside of the vesicles)

fluid mosaic model

• “fluid” indicates that molecules diffuse laterally

• the lateral diffusion, flexion, and rotation (but no flip-flopping) of phospholipids increases the fluidity of the membrane

• cells may decrease the fluidity of the membrane in response to higher temperaturesby increasing cholesterol concentration

• “mosaic” is included to account for the many types of molecules residing in the plasma membrane

amphipathic nature of membrane phospholipids

• the hydrophilic, polar phosphate head is in contact with water/other hydrophilic molecules

• the 2 hydrophobic, nonpolar tails of each phospholipid in the first layer spontaneously arrange themselves so that they turn inwards to contact those in the second layer rather than the external aqueous environment

• membrane separates extracellular fluid from the intracellular cytosol

• the hydrophobic tails are responsible for the membrane’s bilayer structure

spontaneous formation of the phospholipid bilayer

• phospholipid bilayers often form sealed compartments to shield hydrophobic tails from water (since their exposure to water is energetically unfavorable)

• if the bilayer remained planar, both the hydrophilic heads and hydrophobic tails would be exposed to the external aqueous environment

vesicles

• small, membrane-bound sacs within cells that store, transport, or digest cellular products like proteins and lipids

• constructed either by the Golgi apparatus or by the cell pinching in on itself and fusing, trapping a substance within it

phospholipid structure

• glycerol acts as the three-carbon backbone to which fatty acids and phosphate are attached

• glycerol is connected to two hydrophobic fatty acid tails at carbons 1 and 2, and a hydrophilic phosphate group at carbon 3

• each tail contains 14-24 carbon atoms, with one containing a double bond that kinks the tail to create more space between lipid molecules

• in sum, polar heads are generally comprised of a phosphate group and a small organic molecule (e.g., choline or serine) that increases the hydrophilicity of the head

• phosphatidylcholine is the most common phospholipid, with choline being attached to the phosphate group as the top-most structure of the polar head

unsaturated hydrocarbons

• have a double bond between two carbon atoms that creates a kink in the chain, causing the hydrocarbon to bend

• contrasts with saturated hydrocarbons that remain linear

• in nature, hydrogens are always cis (on the same side of the carbon atoms engaged in the double bond), enabling them to maintain membrane fluidity; in contrast, trans hydrocarbons have straighter structures that allow them to pack more tightly together

• the kink in one chain adds to the fluidity of the membrane since the other atoms are free to rotate around this double bond

membrane phospholipid movement

• phospholipids can flex, rotate, and diffuse but rarely flip flop (as in the two layers don’t switch)

• flexion: fatty acid tails wiggle and bend, increasing space between molecules to prevent the membrane from becoming too rigid or freezing at low temperatures

• rotation: can spin around their own axis, allowing them to adjust their position without moving from specific location

• diffusion: move sideways, often exchanging positions with other phospholipids in their own monolayer

• do not flip flop: incapable of moving from one monolayer of the bilayer to the other since the hydrophilic head would have to pass through the hydrophobic core of the membrane to do so

how does the phospholipid bilayer respond to increases in temperature?

• higher temperatures mean faster rate of diffusion, and the membrane is more fluid

• cells insert phospholipid molecules with longer tails or fewer double bonds, and cholesterol in response to higher temperature

• longer tails move more slowly, making the membrane more viscous to prevent the membrane from becoming leaky

• increased saturation of phospholipid molecules means the tails pack together more closely, decreasing space between molecules and fluidity

• cholesterol fills spaces between tails, widening and stiffening the bilayer

where do basic phospholipid membranes come from?

• phospholipid precursors are assembled into the bilayer on the cytosolic surface of the smooth ER (not the luminal side)

• new lipids are initially only added to the cytosolic half of the smooth ER lipid bilayer, creating an initial imbalance

• once produced, the ATP-independent Scramblase enzyme embedded within the ER membrane facilitates the random and equal distribution of phospholipids between the inner (cytosolic) and outer (luminal) leaflets of cell membrane

• results in symmetric growth of both halves of the bilayer

• phospholipids are then shipped to the Golgi apparatus

Golgi apparatus membrane assembly

• phospholipid bilayers synthesized with the help of ER Scramblases are first shipped to the Golgi apparatus

• the bilayers the Golgi receives are fairly symmetrical

• ATP-dependent transmembrane enzymes called “flippases” establish membrane asymmetry by selectively moving specific phospholipids from the lumenal side to the cytosolic side

• this asymmetry curves the membrane by creating disparities between the inner and outer leaflets

Golgi apparatus lipid transport to plasma membrane

• Golgi-derived vesicles can fuse with the plasma membrane

• upon fusion with the plasma membrane, the orientation of the Golgi membrane and associated proteins is maintained (i.e., the luminal side becomes the extracellular side of the plasma membrane, and the cytoplasmic side of the Golgi membrane remains facing the cytoplasm)

• addition of sugars to lipids and proteins (called glycosylation) primarily occurs in the Golgi lumen

lumen vs cytosol

• lumens are the internal space of an organelle

• the lumen has a high concentration of calcium and specialized enzymes

• cytosol is the aqueous fluid surrounding organelles within a cell

• the cytosol contains metabolic enzymes, ribosomes and the cytoskeleton

distribution of phospholipids at the cell’s plasma membrane

• phospholipid heads differ between extracellular and cytosolic cides

• phosphatidylcholine and sphingomyelin are highly concentrated on the extracellular side

• phosphatidylserine and phosphatidylethanolamine are highly concentrated on the cytosolic side

• flippases are responsible for the differential distribution of phospholipids at the plasma membrane

• glycolipids are only found in the outer, extracellular leaflet, added to lipids in the Golgi lumen

significance of phospholipid modifications

• specific modifications act as signaling cues, particularly for apoptosis and immune recognition

• for example, during apoptosis phosphatidylserine flipds form the inner to outer leaflet

• is why flippases are really important (ensure the specific phospholipids are on the correct side of the membrane)

• modified phospholipids create binding sites for the signaling domains on glycosylated proteins (i.e., those that have a sugar molecule attached to them)

plasma membrane carbohydrates

• lipids and oligosaccharides (composed of less than 15 sugar units) form glycolipids

• proteins and oligosaccharides (composed of less than 15 sugar units) form glycoproteins

• proteins and longer polysaccharides form proteoglycans

• these sugars are added to lipids and proteins in the Golgi lumen

• glycoproteins, glycolipids, and proteoglycans join to form the glycocalyx on the external surface of the plasma membrane

• the glycocalyx occurs only on the external surface of the plasma membrane, where it protects the surface and serves in cell-cell recognition (e.g., gamete recognition)

plasma membrane proteins

• proteins embedded within and attached to the cell membrane make up 55% of the cell membrane’s mass

• integral proteins are interwoven and embedded through the width (both layers) of the membrane, attaching to each other and to the extracellular matrix to form channels, transporters, and receptors

• peripheral membrane proteins are bound to either the inner or outer surface of the plasma membrane via weak bonds, enabling them to be removed by gentle procedures that leave the bilayer intact

transport across artificial lipid bilayers and membranes

• small, nonpolar molecules readily diffuse across lipid bilayers (e.g., O2 and CO2)

• larger uncharged polar molecules like glucose diffuse very slowly across artificial lipid bilayers; in membranes, most glucose transport occurs via transporter molecules

gated ion channels

• integral protein channels that switch between open and closed conformations

• gated ion channels vary in the conditions that influenec their opening and closing, enabling them to respond to different types of stimuli

• types include voltage-gated, ligand-gated, and stress-activated

voltage-gated ion channels

• integral protein channels that open and close due to variations in membrane potential

• are highly ion-specific channels that allow particular ions to pass

• controlled by extremely sensitive voltage sensors (specialized charged protein domains on ion channel proteins)

• changes in membrane potential above a certain threshold value exert sufficient electrical force on voltage sensors to encourage channel to open or close

ligand-gated ion channels

• integral protein channels that open a central pore to allow specific ions like Na+ or K+ to flow across the membrane in respose to binding of a ligand to the channel protein receptor

• the binding of either extracellular or intracellular ligands induces a rapid conformational change which opens the gate, creating a pathway for rapid passive ion movement

• examples: GABA, acetylcholine, glycine

stress-activated channels

• integral protein channels that convert physical forces into electrical or chemical signals

• allow ions to flow across the membrane in response to physical deformations that trigger changes in the bilayer, cytoskeleton, or extracellular matrix

• example: sounds triggering the opening of ion channels in ear hair cells

how are stress-gated ion channels involved in hearing?

• the basilar membrane below and the tectorical membrane above are sheets of extracellular matrix that sandwich the organ of Corti to enable hearing

• sound vibrations cause the basilar membrane to vibrate up and down, causing stereocilia to tilt

• the bending of stereocilium stretches the fine filaments that link them to the next shorter filament, pulling open stress-gated ion channels in the stereocilium membrane, allowing positively charged ions to enter from the surrounding fluid

• influx of positive ions activates the hair cells, which stimulate underlying endings of the auditory nerve fibers that convey the auditory signal to the brain

structural classification of membrane proteins

• integral proteins are permanently embedded within the lipid bilayer and are most often transmembrane

• integral includes transmembrane, monolayer associated, and lipid-linked

• peripheral proteins are those that temporarily associate with the membrane’s surface or integral proteins

• peripheral includes protein-attached proteins

integral membrane proteins

• proteins that are interwoven and embedded through both layers of the membrane

• used for membrane transport, serve as receptors (glycoproteins) and enzymes (to a lesser extent)

• transmembrane proteins: cross the entire lipid bilayer

• monolayer-associated proteins: embedded in only one leaflet of the bilayer

• lipid-linked: covalently attached to a lipid anchor that inserts itself into the membrane

functional classifcation of membrane proteins

• leak channels, ion gated channels, and porins all fall under the transmembrane channel protein category

• transporters (carrier) are also transmembrane proteins

• protein-attached proteins are peripheral

channel proteins

• integral proteins that have a hydrophilic pore

• are selective for specific solutes

• allow for faster transfer of solutes between the inside and outside of the cell

• leak channels are integral proteins that are always open, allowing H2O and ions to move due to concentration gradients

• always function in passive transport

transporter proteins

• integral membrane proteins that function by binding a solute and undergoing a conformational change that enables the movement of solutes across the membrane

• function in either passive or active transport depending on type (uniport, symporters, and antiporters)

• two conformational states: (1) binding sites exposed to extracellular space (2) binding sites exposed to cytosol

• generally speaking, substrate affinity is high in its extracellular facing conformation, and low when in its cytosolic facing conformation to release substrate into the cell

types of transporters/pumps

• uniport: transports only one solute with no direct coupling to another solute; the substrate binds, protein changes shape, and the substrate is released on the other side

• symporters: moves two or more different solutes in the same direction by coupling the transport of one solute to the transport of another; generally use theenergy released via passive transport of one to power active transport of the second

• antiporters: solutes are transferred in the opposite direction by using the nergy derived from the downhill transport of one to pump another molecule up its gradient

types of transport

• simple diffusion: the movement of small, uncharged, NP and lipid-soluble molecules from areas of high to low concentration through cell membranes

• passive transport: facilitated diffusion of larger or polar molecules utilizing either transporters or channel proteins; does not require ATP but does need kinetic energy for movement

• active transport: the ATP-dependent pumping of substances against the concentration gradient (from areas of low to high concentration) using specific transporter (pump) proteins

passive vs active transport

• passive transport does not require an input of energy as molecules move down the concentration gradient from areas of high to low concentration

• primary active transport involves the direct use of an energy source (either light, redox energy, or ATP hydrolysis) to move molecules against a concentration gradient

• secondary active transport utilizes existing electrochemical gradients established by primary active transport to drive the movement of other substances

electrochemical gradients

• when a charged particle moves across a membrane, its movement is determined both by the chemical and electrical gradient

• the chemical gradient drives particles from high to low concentrations

• the electrical gradient is driven by opposite charges attracting and like chargs repelling

• if both work in the same direction, the ion moves readily with large net movement

• if they work in opposite directions, net movement depends on size of concentration difference and magnitude of the membrane potential; if one is stronger, movement occurs in that direction

establishment and maintenance of electrochemical gradients

• electrochemical gradients=concentraiton gradient of ions and electrical charge across the membrane

• the cytosolic side of the membrane is negatively charged due to the accumulation of negatively charged molecules like phospholipids, proteins, and phosphates

• when positive ions accumulate outside the cell, membrane potential increases the movement of positive ions that can readily enter due to both concentration and charge

• when positive ions accumulate inside the cell, ions move due to concentration gradient but are slowed by membrane potential (electrochemical gradient is not as large)

membrane potential

• the electrical potential difference between the outer and inner sides of the cell’s plasma membrane due to a slight excess of positive ions on one side and negative ions on the other

• in cells, membrane potential is between -80 and -40 mV

glucose transport

• glucose is too large to pass via simple diffusion

• the glucose uniport is a transmembrane protein that switches between different conformations

• the uniport binds glucose on one side, undergoes a conformational change, and releases it on the other side

• when blood glucose is high, glucose binds to the extracellular binding site to be released into the cell (and opposite when blood glucose is low)

• glucose is electrically uncharged, so concentration gradient alone drives this passive transport

transporter-mediated active transport

• gradient driven pump: the movement of one ion down its gradient releases energy that is then used to move another ion up its gradient, coupling their movement

• ATP-driven pump: the binding and hydrolysis of ATP causes a conformational change that moves solutes against their concentration gradient (e.g., lysosome proton pumps)

• light-driven pump: conformational changes in pigment molecules bound to pumps acts as a stressor that causes the protein pump itself to change shape and pump ions against their concentration gradient (e.g., bacteriorhodopsin pumping protons against an electrochemical gradient)

glucose-sodium ion symport pump

• a secondary active transport proccess that moves glucose into cells against its concentration gradient by coupling it with the downhill movement of sodum ions from high concentration in the extracellular space to a low concentration in the cytoplasm

• upon binding two sodium ions, the transporter moves into an outward open conformation, allowing glucose to bind

• once both have bound the transporter, it changes into its occluded-occupied state, trapping the substrates in the membrane

• the pump then changes to its inward-open state, releasing sodium and glucose into the cytoplasm

• the now-empty transporter returns to the outward facing conformation to begin again

• enables continual absorbption of glucose even once intracellular glucose concentration becomes relatively high

sodium-potassium antiport pump

• an antiport pump that uses ATP to maintain homeostasis (low intracellular sodium and high intracellular potassium) in a matter of 10 milliseconds per cycle

• sodium ions bind on the cytosolic side of the pump

• ATP hydrolysis then changes conformation of the Na/K ATPase, pumping out 3 Na

• this conformational change also enables binding of K ions, triggering the release of the phosphate group and pumping in 2 K ions

• high extracellular concentration of Na needed since its import is coupled to movement of other molecules across membrane (e.g., glucose-sodium symport)

• high cytoplasmic K concentration needed for proper functioning of nervous system

gut epithelial cell membrane transport

• glucose-sodium symport transporters are used in scenarios where two solutes are needed for the effective uptake of a certain nutrient (e.g., both glucose and sodium are needed in oral rehydration therapy to replace sodium loss)

• glucose absorption occurs in the apical domain of the plasma membrane via secondary active transport relying on the glucose-sodium symport

• the basolateral domain of the membrane contains high concentration of Na/K antiport pumps that maintain the low intracellular sodium concentration that drives the import of glucose

• glucose also leaves the cell to enter the bloodstream via faciliated diffusion through uniport transporters located on the basolateral membrane

• all three types of transporters are maintained in separate domains (apical surface vs basolateral surface) by tight junctions that seal adjacent epithelial cells

transporter/pump summary

• plasma membrane transporters provide nutrientsand maintain homeostasis (e.g., maintain low intracellular Na and high intracellular K)

• perform both passive and active transport

• can use transporters and pumps to move ions, glucose, and other solutes through membrane

• passive transport driven by concentration gradient

• active transport use different concentration gradients between two solutes or energy provided by ATP or light

• three types of transporters: uniporters, symporters, and antiporters

osmosis

• a type of channel-mediated passive transport in which water diffuses across a membrane from areas of high to low concentration

• aquaporin channels are the primary conduit for water moving across membrane, allowing our kidneys to reabsorb 99% of water

• in a hypotonic solution where there is a higher intracellular concentration of solute as compared to the extracellular environment, water will rush in, causing the cell to swell until the membrane ruptures

• in hypertonic solutions where there is a higher extracellular solute concentration, water rushes out of the cell, the cell loses volume, and the membrane beomes wrinkled or shriveled (crenated)

action potentials

• is an electrical discharge (movement of charge) triggered by a rapid change in membrane potential that enables communication between neurons

• driven by the opening of voltage-gated Na channels: when the dendrites of one neuron receive neurotransmitters from another neuron, a few Na+ channels open on a small segment of the axon, causing positive ions to diffuse through the channel

• this increases the concentration of positive ions in that segment of the axon, raising the intracellular charge to a less negative state (depolarization)

• depolarization to about -40mV causes adjacent Na+ channels to open, and the action potential propagates along the length of the axon

• action potentials are fired in an all or nothing manner, meaning neurons can only produce a full-strength signal so long as threshold potential is reached

repolarization

• as soon as the intracellular charge along a segment of the axon reaches +40mV, the Na channels close/inactive to prevent further Na+ influx, terminating the action potential

• voltage-gated K+ channels then open, and K+ ions exit the cell

• with no additional Na+ ions entering the cell and K+ ions leaving the cell, the intracellular charge becomes more negative, eventually reaching a more negative state than its resting potential

• Na+ and K+ concentrations are then returned to resting potential levels via Na+/K+ antiport pumps

• the membrane potential then becomes more negative than resting (is hyperpolarized) due to the slow closing of K+ channels

changes in neuronal membrane potential

• resting membrane potential is around -60mV

• voltage-gated Na+ channels are closed between -80mV and -40mV

• a stimulus depolarizes the plasma membrane until the membrane potential is raised to about -40mV

• once the membrane potential reaches -40mV (threshhold), an action potential is triggered

• after an action potential is triggered, the membrane rapidly depolarizes and reaches +40mV before it returns to its resting negative value

ion channel selectivity

• most ion channels have a selectivity filter size that is ideal for their respective ions

• the selectively filter is the narrowest part of the channel, with its diameter and shape enabling their respective ions to fit perfectly

• amino acids in the interior of the pump interact with the ion to stabilize the ion as it squeezes through

• these channels that are selective specific ions are involved only in passive transport

• are responsible for maintenance of membrane potential

resting membrane potential

• refers to the concentration of ions on each side of the plasma membrane when no neural impulse (action potential) is traveling through the neuron

• in a resting neuron, there is a high intracellular concentration of K+ and a high extracellular concentration of Na+ since due to Na/K pumps pumping out 3 Na+ ions for every 2 K+ ions it pumps in (more positive charge leaves the cell than enters it)

• the ions are arranged in a thin layer (smaller than 1 nm) next to the membrane

• despite there being no impulse traveling through the neuron, the cell remains charged since there are unequal concentrations of positive and negative ions on each side of the plasma membrane

• the resting membrane potential is about -60 mV since large, negatively charged proteins and Cl anions outnumber the concentration of intracellular K+ (especially since K+ ions continuously leak out through membrane channels)

• the resting neuron is said to be polarized since there is a positive charge on one side of the plasma membrane and a negative charge on the other

synapses

• neurons transmit chemical signals across synapses, which are composed of three parts: presynaptic (sending) neuron, synaptic cleft, postsynaptic (receiving) neuron

• both presynaptic and postsynaptic membranes are thickened at the synapse, reflecting areas where specialized proteins and chanels are concentrated to enable efficient signal transmission

• when an action potential reaches a nerve terminal of the presynaptic neuron, the membrane depolarizes to open voltage-gated Ca2+ channels in the terminal membrane (which are closed at rest)

• because the concentration of Ca2+ is much higher outside the neuron than inside, Ca2+ rapidly flows into the nerve terminal

• increased Ca2+ in the nerve terminal stimulates the synaptic vesicles to fuse with the plasma membrane and exocytose their neurotransmitters into the synaptic cleft

• the released neurotransmitter diffuses across the cleft to bind to and open transmitter-gated ion channels in the plasma membrane of the postsynaptic cell

• once these channels open, ion movement creates a new electrical signal in the postsynaptic cell and may trigger an action potential if the depolarization reaches threshold (converts the chemical signal back into an electrical one)

channel proteins and membrane potential summary

• ion-selective channels have a filter that allows for the passage of only channel specific proteins

• leakage of K+ through its ion channels and the Na+/K+ antiport pump make the plasma membrane have a relative negative charge

• beacuse ions line up in a thin layer (<1nm) right next to the membrane, relatively small changes in the number of ions can drastically alter the membrane potential

• the positioning of ions, permeability of membranes, and voltage-gated channels make cells electrically excitable

• action potentials occur when voltage-gated Na+ channels open rapidly and in sequence down neuronal axons

• membrane resting potential is reestablished by the closing of voltage-gated Na+ channels together with the opening of K+ channels and ATP-dependent Na+/K+ antiport pumps

food energy

• stored in chemical bonds within food molecules

• most important fuel sources are sugars

cell metabolism

• the sum of all chemical reactions occuring in a cell and within an organism

• there are two types: anabolism and catabolism

anabolism

• endergonic reactions that utilize energy (from catabolism) to build more complex molecules and structures that the organism needs from simple precursors

• example: protein synthesis

catabolism

• the exergonic decomposition/breakdown of molecules to release energy from their chemical bonds

• these reactions are most often associated with the generation of adenosine triphosphate and carrier molecules such as nicotinamide adenine dinucleotide (NADH)

• also refers to any reaction where a complex molecule is broken into a smaller one (e.g., to release the energy stored in food)

waste generation during cellular metabolism

1. decomposition of food molecules: more complex macromolecules must be broken down extracellularly before they can enter your cellls; although occasionally, lysosomes can digest large molecules in the cell’s interior; produces intermediate wastes (e.g., monosaccharides, fatty acids and glycerol, amino acids, etc.)

2. citric acid cycle: produces carbon dioxide and water as byproducts (cellular waste); occurs mainly in the cytosol, except for the final step in which pyruvate is converted to acetyl groups on acetyl CoA (which occurs in the mitochondria)

3. complete acetyl CoA oxidation occurs entirely in the mitochondria, proucing CO2 waste

sugar oxidation

• controlled oxidation of sugar in the cell preserves energy by enabling storage in ATP rather than releasing it all simultaneously as heat

• enzymes catalyze oxidations via a series of small steps in which free energy is transferred to carrier molecules (usually ATP and NADH)

• at each step, enzymes overcome small activation energy barriers at cell temperature so that reaction can occur

• total free energy released by controlled oxidation of glucose in the cell is exactly the same as that released as heat by direct burning of sugar in a nonliving system; it’s just more directly utilizable when controlled

glycolysis

• the first set of reactions for both aerobic cellular respiration and anaerobic fermentation

• uses carbohydrates to generate precursors for the TSA and ETC

• divided into two 5-step phases: energy investment and energy payoff

• net products: 2 pyruvate molecules, 2 ATP (2 used as activation energy and 4 released), 2 NADH

energy investment phase of glycolysis

1. first phosphorylation: hexokinase phosphorylates the oxygen on carbon 6 to make glucose-6-phosphate; the polar phosphate group traps the molecule within the cell, and reduces concentration of pure glucose enabling more to enter cell via diffusion (costs 1 ATP)

2. isomerization: phosphoglucose isomerase catalyzes the isomerization of G6P into fructose-6-phosphate (F6P)

3. second phosphorylation: phosphofructokinase adds another phosphate to the carbon 1 hydroxyl to create fructose-1,6-bisphosphate using a second ATP

4. cleavage: fructose bisphosphate aldolase utilizes the energy from the 1st and 2nd phosphorylations to cleave fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP)

5. isomerization: triosephosphate isomerase converts DHAP into a second molecule of GAP

energy payoff phase of glycolysis

6. oxidation: glyceraldehyde phosphate dehydrogenase uses an inorganic phosphate to convert the two GAP molecules into 1,3-bisphosphoglycerate and reduces NAD+ to NADH

7. first ATP generation: phosphoglycerate kinase catalyzes the transfer of a phosphate group to ADP, converting 1,3-bisphosphoglycerate to 3-phosphoglycerate and producing 1 ATP per GAP molecule (2 ATP total)

8. phosphate transfer: phosphoglycerate mutase isomerizes 3-phosphoglycerate into 2-phosphoglycerate

9. dehydration: enolase catalyzes the loss of a hydroxyl group, producing phosphoenolpyruvate (PEP)

10. second ATP generation: pyruvate kinase transfers the remaining phosphate group to ADP, generating another ATP and converting PEP into pyruvate

net ATP generation during glycolysis

• 2 ATP must be used to provide the energy needed to initiation glycolysis since glucose alone is not a reactive molecule

• one ATP is used in step 1 of glycolysis, and a second is used in step 3

• four total ATP are generated during the second half of glycolysis as GAP is being converted to pyruvate

• all ATP synthesis during glycolysis is done via substrate-level phosphorylation as it involves the transfer of a phosphate group directly from a substrate molecule

reversible reactions of glycolysis

• all except three steps are reversible, with the irreversible steps being highly exergonic (it would take too much energy to reverse these)

• the reveribe steps all operate near equilibrium, meaning the concentrations of involved molecules enable the reaction to proceed in either direction depending on cellular environment

• first phosphorylation of glucose by hexokinase=irreversible

• second phosphoryation of fructose-6-phosphate by phophofructoinase=irreversible

• transfer of phosphate group from phosphoenolpyruvte to ADP=irreversible

anaerobic cellular respiration

• process by which energy within the chemical bonds of foodstuffs are transferred to a form (ATP) that the cell can utilize without oxygen

• occurs in the cytoplasm and results in the very rapid generation of ATP, in about 8 seconds

• only get 2 net ATP molecules per glucose molecule

• two pathways: lactic acid fermentation and alcoholic fermentation

lactic acid fermentation

• anaerobic cellular respiration pathway in which pyruvate produced by glycolysis is converted to lactate

• in the absence of oxygen, pyruvate oxidizes NADH which converts the pyruvate into lactate, regenerating NAD

• purpose is to regenerate NAD+ from NADH to ensure there is NAD+ available for glycolysis to continue producing 2 net ATP

• yields much less overall energy than complete oxidation (36-38 ATP in aerobic respiration vs 2 in lactic acid fermentation)

• often occurs in muscle cells during intense exercie or in microorganisms like lactic acid bacteria

alcoholic fermentation

• occurs in some organisms that grow anaerobically (e.g., yeast)

• pyruvate is converted into acetaldehyde by pyruvate decarboxylase

• acetaldehyde is then reduced to ethanol, regenerating NAD+ from NDH for continued glycolysis

citric acid cycle (TCA)

• occurs in the mitochondrial matrix

• the TCA alone is anaerobic but its products are necessary for aerobic ETC

• energy released in TCA reactions are stored in NADH and FADH2

• is an 8 step process: citrate formation, isomerization, first oxidative decarboxylation, second oxidative decarboxylation, substrate-level phosphorylation, dehydrogenation, hydration, oxidation

• acetyl CoA must be formed prior so that it can be used in initiating the cycle (via pyruvate oxidation or beta oxidation of fatty acids)

• net products: 3 NADH, 1 FADH2, 1 GTP, 2 CO2

diffusion of pyruvate into the mitochondrial matrix

• pyruvate crosses the outer mitochondrial membrane to enter the intermembrane space via passive diffusion through mitochondrial porins

• once inside the intermembrane space, pyruvate crosses the inner mitochondrial membrane via the mitochondrial pyruvate carrier

lipolysis and beta oxidation

• stored triglycerides are broken down into glycerol and free fatty acids via lipolysis

• glycerol generally enters the bloodstream for transport to the liver to enter either gluconeogenesis or glycolysis

• fatty acids are transported to the mitochondrial matrix where enzymes remove 2-carbon units via a four step cyclic process that results in the production of acetyl-C

• the reactions trim two carbons at a time from the carboxyl end of the fatty acids, continuing until the fatty acid is completely degraded

• each turn of the beta oxidation cycle produces 1 NADH, 1 FADH2, and 1 acetyl CoA molecule

utilization of intermediate molecules synthesized during glycolysis and TCA

• intermediates are molecules that link reactions but do not go back and forth between stages (i.e., products of one reaction that are used as inputs for the next reaction)

• many of the intermediates formed in glycolysis and TCA are used for biosynthetic (anabolic) pathways, where they are converted into amino acids, nucleotides, lipids, and other small organic molecules

glycolysis intermediate biosynthetic pathways

• the intermediate molecules are essential precursors for the synthesis of other biological molecules

• glucose-6-phosphate=nucleotides

• fructose-6-phosphate=amino sugars, glycolipids, glycoproteins

• dihydroxyacetone phosphate=lipids

• 3-phosphoglycerate=serine

• phophoenolpyruvate=amino acids, pyrimidines,

• pyruvate=alanine

TCA intermediate biosynthetic pathways

• citrate=cholesterol, fatty acids

• alpha-ketoglutarate=glutamate, other amino acids, purines

• succinyl CoA=heme, chlorophyll

• oxaloacetate=aspartate, other amino acids, purines, pyrimidines

gluconeogenesis overview

• broadly speaking, “reverses” step 3 of glycolysis (the production of fructose-1,6-bisphosphate via phosphofructokinase)

• is the process by which blood glucose is synthesized from small non-carbohydrate organic molecules like lactate (from fermentation; first converted into pyruvate), pyruvate, amino acids, glycerol (from lipolysis)

• the production of 1 glucose molecule requires 4 ATP, 2 GTP, and 2 NADH so it can only be done when the liver has plenty of energy in the form of pyruvate, citrate, and/or ATP

• gluconeogenesis (the reverse of step 3) is regulated by fructose-1,6-bisphosphatase

control point of gluconeogenesis

• the major control point of glycolysis is step 3, in which glucose is broken down to synthesize fructose 1,6 bisphosphate

• once the fructose-1,6-bisphosphate is made, the cell has essentially committed to continuing glycolysis

• phosphofructokinase is activated by AMP, ADP, and inorganic phosphate (the byproducts of ATP hydrolysis), and inhibited by ATP, citrate, and fatty acids

• when energy reserves (ATP) are low, there are high concentrations of AMP and ADP (non-phosphorylated ATP precursors) that activate glycolysis

• when energy reserves (ATP) are high, ATP, citrate, and pyruvate accumulate; since the liver doesn’t need to produce more ATP via glycolysis, it begins synthesizing glucose via gluconeogenesis

• fructose 1,6-bisphosphatase regulates the reverse reaction and is inhibited by byproducts of ATP hydrolysis and activated by ATP, citrate, and fatty acids

glycogen storage

• with an excess of glucose, animals store glycogen (a branched polymer of glucose) to provide energy during fasting

• glycogen is large and surrounds itself with lots of water, so it occupies lots of space in cells

• in glycogen, carbon 1 of one glucose bonds to carbon 4 of another; once incorporated, each gluucose unit is called a glucose residue

• branch points occur when carbon 1 of one residue bonds to carbon 6 of another, creating new chains

• glycogen forms granules in the cytoplasm of a liver cell

• when more ATP is needed than can be generated from food molecules taken from the bloodstream, cells break down glycogen

glycogen conversion to glucose (glycogenolysis)

• when more ATP is needed than can be generated from food molecules in bloodstream, cells break down glycogen to supply glucose

• glycogen phosphorylase cleaves 1-4 glycosidic bonds and adds an inorganic phosphate to produce glucose-1-phosphate

• phosphoglucomutase then converts glucose-1-phosphate into the glucose-6-phosphate molecule that enters glycolysis to generate ATP

• the synthesis and degradation of glycogen are both regulated by glucose-6-phosphate, but in opposite directions

• glucose-6-phosphase activates synthesis but inhibits glycogen breakdown by inhibiting glycogen phosphorylase activity (along with ATP)

fat droplets vs glycogen stores

• fats are far more important for storing energy than glycogen since the oxidation of a single gram of fat releases ~2 times as much energy as oxidation of a gram of glycogen

• glycogen binds a great deal of water, producing a six-fold difference in actual mass of glycogen required to store same amount of energy as fat

• on average, adult humans store enough glycogen for only about a day of normal activity, but enough fat to last nearly a month

• fat droplets accumulate in adipose cells

eukaryotic vs prokaryotic cells

• both have: cell membrane, cytoplasm, and DNA

• eukaryotic cells have organelles and are much more advanced than prokaryotic cells

• prokaryotic cells don’t have a nucleus, mitochondria, or chloroplasts

mitochondria vs chloroplasts

• both are present only in eukaryotic cells

• both are involved in converting energy from one form to another; mithochondria convert chemical energy stored in foodstuffs to chemical energy stored in ATP, while chloroplasts convert between light energy and chemical energy stored in ATP

• both have multiple layers of membranes filled with enzymes

how are mitochondria and chloroplasts similar to bacteria?

• all three contain their own set of DNA, are able to replicate independently, and make RNA and proteins

• both store genetic material as circular DNA with sequences that resemble that of bacteria

• DNA within the two organelles lack histones, and their ribosomes resemble thos ein bacteria, both in terms of size and in their sensitivity to antibiotics

• mitochondria and chloroplasts undergo fission to replicate, much like bacteria

mitochondrial DNA

• mitochondria contain their own genome which is about 16,000 base pairs long

• stored within the mitochondrial matrix, locoated inside the mitochondria organelles, which are situated in the cell’s cytoplasm

• in humans, mitochondrial DNA is inherited exclusively from the mother since, during fertilization, paternal mitochondria from the sperm are destroyed, leaving behind only the egg’s; has applications in tracing maternal lineages

• 13 proteins key to respiratory complexes are encoded by human mitochondrial DNA

mitochondria

• are found in all eukaryotic cells (not prokaryotes) as a defining feature (e.g., animals, plants, fungi)

• are derived from and resemble organisms of the bacterium domain

• can replicate independently of the cell to create more ATP

• have an external phospholipid bilayer (mitochondrial membrane) and a folded (to increase surface area) internal phospholipid bilayer called the cristae (which is embedded with thousands of proteins)

• have two compartments: intermembrane space and matrix

• its chemical composition gives it an orange appearance

• comprises ~22% of total cell volume, with about 1700 per cell

mitochondria adaptation

• mitochondria adapt to meet energy demands, with size being relative to the cell and its energy needs

• mostly located in the cytosol where energy demand is highest

• number increase with increasing energy demand (e.g., in skeletal muscles as individuals train to run long distances, cardiomyocytes, sperm cells)

• in some cells, are large and have several branches to form long, interconnected tubular networks

mitochondrial dynamicism

• mitochondria are incredibly dynamic, spontaneously undergoing fission and fusion

• fluorescent micrographs taken at 3 mintue intervals show the dynamic nature of the mitochondria, with mitochondria moving alone cytoskeletal tracks, changing shape, fusing together, splitting apart, etc.

• GTP powers the motor molecule dynamin to divide a mitochondrion via fission

• dynamic behavior important for distributing ATP, removing damaged sections, responding to metabolic stress, maintaining mitochondrial DNA integrity

mitochondrial fusion

• involves joining together of two mitochondrion to form one larger mitochondrion

• fusion occurs in two steps: (1) outer membrane fusion by mitofusins 1 and 2, and (2) inner membrane fusion by OPA1

• the proteins utilized in fusion are GTPases that use GTP for energy to change shape and pull membranes together

• fusion mixes mitochondrial contents, dilutes damaged components, helps maintain ATP production, and supports survival during stres

• enables a slightly damaged mitochondrion to be functionally rescued by fusion since the two mitochondrion are able to share contents

mitochondrial fission

• fission refers to splitting of one mitochondrion into two smaller mitochondria, allowing it to divide in a manner similar to bacteria

• dynamin related protein 1 is recruited to the mitochondrion’s outer membrane, forming a ring around the mitochondrion

• DRP1 hydrolyzes GTP, tightening the ring and constricting the membrane to the point that the mitochondrion divides

• helps distribute mitochondria during cell division, allows transport of mitochondria to areas needing ATP, isolates damaged sections, participates in apoptosis

dynamin

• GTPases that use guanosine triphosphate hydrolysis as an energy source

• dynamin wraps around a constriction site on the mitochondrion

• GTP hydrolysis causes a conformational change in dynamin, constricting the mitochondrial membrane

• this constriction splits the mitochondrion in two during fission

• also essential for the pinching off of clathrin-coated endocytic vesicles from plasma membranes

mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS)

• a mitochondrial disease caused by mutations in several mitochondrial genes

• affects brain and muscles

myoclonic epilepsy with ragged-red fibers (MERRF)

• a mitochondrial diseas caused by mutations in several mitochondrial genes

• affects the brain and muscles

leigh syndrome

• caused by mutations in several mitochondrial genes

• affects brain and spinal cord