Exam 3 Biology

Hexokinase

The enzymes that catalyzes the phosphorylation of glucose to form glucose-6-phosphate in the first step of glycolysis HK structure without glucose (open, looks concave); with glucose (closed) This reaction is essentially irreversible - energetically unfavorable to proceed in reverse direction (Gly-1 is a commitment to glycolysis) regulates entry of glucose into glycolysis

Warburg effect

-use of glycolysis under normal oxygen conditions (aerobic glycolysis) -allows products of glycolysis to be used for rapid cell growth -activated by oncogenes and mutant tumor suppressors

Energy investment steps in glycolysis

1 and 3 - ATP needed to add phosphate group these steps are highly exergonic

Catabolism vs Anabolism

Catabolism is oxidative and exergonic, anabolism is reductive and endergonic Catabolism (breakdown) and Anabolism (build up) catabolism: break down of organic molecules anabolism: synthesis of organic molecules

Enzymes

Catalysts for chemical reactions in living things decrease the activation energy (Ea) required for a chemical reaction to proceed CANNOT change reaction equilibrium CANNOT alter ΔG to make its reaction thermodynamically possible

active site

The part of an enzyme or antibody where the chemical reaction occurs.

energy transduction

Pigments undergo photoexcitation - pigments absorb particular wavelengths and quantities of light energy

3 ways an enzyme lowers activation energy (Ea)

Positioning substrate(s) Making an 'attack' on bonds Stabilizing the transition state (ES)

cellular respiration

Producing ATP with oxygen by breaking down glucose (36-38 ATP); occurs in the mitochondria

allosteric inhibition

The process in which an enzyme's activity is changed because of binding to the allosteric site.

Pyruvate

Step 10- (last step) end product of glycolysis influenced by early status of glycolysis, ATP levels, & citric acid cycle status

Phosphofructokinase-1 (PFK-1)

Step 3- regulated enzyme of glycolysis Reactants-- fructose-6-phosphate & ATP- Products-- fructose-1,6 -bisphosphate & ADP also irreversible

energy payoff steps of glycolysis

Step 6, 7, and 10 4 ATP are formed 2 NADH and 2H+ are formed 2 Pyruvate and 2H2O are formed

Km

Substrate concentration at 1/2 Vmax (half way up the reaction line on graph) is the Michaelis-Menten constant (aka Michaelis constant) Km is a measure of the affinity of enzyme for substrate! (

T/F-- Cells couple endergonic reactions to compensating exergonic reactions getting an overall exergonic reaction

True

T/F Cellular machinery (enzyme based) can couple an energetically favorable reactions to an energetically unfavorable reactions making an overall reaction that is energetically favorable.

True- favorable always beats unfavorable

EA

activation energy - the energy required to stabilize the transition state. (Ea) is required for the transition state to be reached in the reaction. (The Ea requirement is the reason we don't see spontaneous reactions taking place immediately.)

metabolism

all the chemical reactions that take place in a cell

amino acid job for the catalytic portion of the active site

amino acids encourage chemical reactions

what can effect the activity of an enzyme?

Temperature, pH, and presence of ions

How is the citric acid cycle regulated?

feedback inhibition (process in which the product or result stops or limits the process....inhibiters binding to the enzyme to stop or slow a reaction)

reversible inhibitors

form weak bonds with enzymes and easily dissociate from them

ΔG

free energy change for a process taking place with reactants and products present under nonstandard conditions. ΔG is related to the standard free energy change

reduction

gain of electrons

cell culture

group of cells grown in a nutrient solution from a single original cell cells grown in vitro (glass/plastic)

Animal cell culture

growth medium needs glucose, amino acids, lipids, vitamins, salts, PH buffer, good CO2 levels, albumin, transferrin, insulin, other hormones, growth factors Used when artificial media are inadequate. Required for growth of viruses and other obligate intracellular parasite.

Vmax

maximum initial velocity or rate of an enzyme-catalysed reaction.

Mouse spleen/mutant cell experiment picture The image shows the hybridoma technology to produce _____________________. Fused cells (step 2) are able to grow on the selective media because they have the ability to grow continually in culture from the ___________________ and have the the ability to synthesize nucleotides from the _____________________. In the last step, cells in each well are tested for their ability to produce antibody with desired characteristics.

monoclonal antibody myeloma cells mouse spleen cells

cultured cells characteristics

monolayer growth cells die (by apoptosis) after 40-60 doublings

inhibitors

decrease enzyme activity

This enzyme's activity ___________ when there is a high concentration of citrate, one of the citric acid cycle products.

decreases

What happens to a reaction with a LOW Km

enzyme has a high affinity for substrate Small Km means enzyme will catalyze reaction efficiently at low substrate concentrations (the reaction takes place with low concentration of substrate)

What happens to a reaction with a HIGH Km

enzyme has a low affinity for substrate Large Km means enzyme requires high concentrations of substrate for efficient catalysis (the reaction won't take place with low concentration of substrate)

Porins

outer mitochondrial membrane channel proteins that allow passage for ions and small particles they are on the outer membranes

coupled reactions

pairs of chemical reactions in which some of the energy released from the breakdown of one compound is used to create a bond in the formation of another compound they have a negative ΔG (free energy) so BOTH can occur

how does electron transport happen in cellular respiration?

sodium electrons flow through complexes I, III, IV

structure of chloroplast

stack of membranes that contain photosythetic pigments (chlorophyll) have 3 membranes

what are the regulated steps of glycolysis?

step 1 (hexokinase) step 3 (phosphofructokinase), step 10 (pyruvate kinase)

Gluconegenesis

synthesis of glucose from noncarbohydrate sources in liver

allosteric activation

the active site becomes available to the substrates when a regulatory molecule binds to a different site on the enzyme

Glycolysis

the breakdown of glucose by enzymes, releasing energy and pyruvic acid. starting glucose reactant to pyruvate product catabolic (breakdown) pathway need to spend 2 ATP and gain 4 ATP AND 2 NADH Involves 10 steps going through 3 phases: 1- preparation and cleavage 2-oxidation and ATP generation 3- pyruvate formation and ATP generation

Michaelis-Menten equation

v = (vmax [S])/(Km + [S]) the mathematical relationship describing the relationship between velocity and concentration - its the kinetics of many enzyme catalyzed reactions NOTE do not need to memorize equation

Coenzymes

vitamins that assist in the normal functioning of enzymes Covalently bound prosthetic groups

Keq

where equilibrium is

Why are metabolic pathways regulated?

without regulation: homeostasis would not be possible counterproductive reactions would occur cells would not be able to respond to their environments no useful cellular energy available

What happens if you have an unregulated enzyme?

would lead to counterproductive energy expenditure and make homeostasis an impossibility

Steps of Glycolysis

1. Phosphate groups from two ATP molecules are transferred to a glucose molecule. 2. A six-carbon is broken down into two three-carbon compounds. 3. Two NADH molecules and four ATP molecules are produced. 4. Each three-carbon compound is converted into a three-carbon pyruvate.

steps in producing a cell culture

1. take tissue sample 2. separate cells from tissues using trypsin/ization 3. grow cells in appropriate media 4. many cells die, some live on to proliferate (make more cells) and are called the primary cell culture 5. When the primary cell culture becomes confluent, a subculture step (aka passage) is done 6. the living cells are called a cell line 7. on a rare occasion the cell may be transformed

ligand

A molecule that binds specifically to another molecule, usually a larger one. bonding is non-covalent some bind weak while other are stronger can alter other bonds

allosteric site

A specific receptor site on some part of an enzyme molecule remote from the active site. binding site other than active site; ligand binding to allosteric site alters conformation

The enzyme is inhibited when the ATP to AMP ratio favors ______________ ATP or AMP

ATP

ATP vs. ADP vs. AMP

ATP has 3 phosphate groups (TRIphosphate) ADP has 2 phosphate groups (DIphosphate) AMP has 1 phosphate group (MONOphosphate)

Non-reversible inhibitors

An inhibitor that binds permanently to the enzyme preventing the enzyme from working.

allosteric regulation

Binding of a small molecule outside the catalytic site Binding may specifically activate or inhibit enzyme activity

How are metabolic pathways regulated?

By controlling when and where enzymes are active. Regulating enzyme activity with allosteric regulation or feedback inhibition

Aerobic cellular respiration in Eukaryotes

Cellular respiration in eukaryotes occurs primarily in the mitochondria. Eukaryotic cells depend on cellular respiration to survive. During glycolysis, the cell breaks down glucose in the cytoplasm with or without oxygen being present. It splits the six-carbon sugar molecule into two, three-carbon pyruvate molecules 1. Glycolysis 2. Pyruvate oxidation 3. Citric acid cycle 4. Electron transport & H+ pumping 5. ATP synthesis cellular respiration is an example of catabolism

cellular respiration vs anaerobic respiration

Cellular respiration that proceeds in the absence of oxygen is anaerobic respiration. Cellular respiration that proceeds in the presence of oxygen is aerobic respiration

confluency

Confluence is the estimated % coverage of cells over the culture surface

How do cells catalyze chemical reactions

Enzymes

Fermentation

Essentially, glycolysis plus additional chemical reaction to recover the NAD+ pyruvate is used to oxidise NADH+ MANY products of fermentation: - Lactic acid - Ethanol- Butyric acid- Acetone- ...More

substrate-level phosphorylation

Formation of ATP with a direct transfer of phosphate from a metabolic intermediate to ADP using a kinase enzyme

what metabolic pathways produce ATP?

Glycolysis citric acid cycle ATP synthesis

anabolic pathways

Metabolic pathways that consume energy to build complicated molecules from simpler ones. ex: photosynthesis

catabolic pathways

Metabolic pathways that release energy by breaking down complex molecules into simpler compounds. Many of the reactions in the pathway are exergonic rxns on their own (-ΔG) ex: cellular respiration

are catabolic and anabolic pathways opposites of each other?

NO, some steps are catabolized by different enzymes and involve different intermediates

metabolic moonlighters

Protein moonlighting is the concept that a gene and its protein can carry out very different roles in a cell We tend to think each protein has a specific role. So recognition of this concept was a bit 'radical'

citric acid cycle

Pyruvate is the product of glycolysis but energy can't be stripped from it any further without it being prepped for the citric acid cycle by being moved and converted to CoA before proceeding to the citric acid cycle The citric acid cycle then completes the breakdown of glucose by oxidizing a derivative of pyruvate to carbon dioxide. Note: 1. pyruvate is moved to the mitochondria and has to be attached to an activated carrier 2. COA is covalently added 3. when it links to COA, NADH is formed (pyruvate is past the membranes by this point)

endergonic reaction

Reaction that absorbs free energy from its surroundings

enzyme active site

Site of the enzyme surface where substrate molecules binds amino acids help bind and activate the substrate (reactant)

Cells are able to conduct chemical reactions 'at the speed of life' by.....

Using enzymes to lower the activation energy of chemical reactions Remember: an enzyme can't change reaction equilibrium (Keq) BUT since cellular concentrations tend to be away from (equilibrium) Keq concentrations, enzymes continue catalyzing reactions towards (equilibrium) Keq An enzyme cannot alter free energy (ΔG) to make its reaction thermodynamically favorable. BUT since cellular reactions strive towards equilibrium enzymes continue using reactions involving free energy to aim for Keq some enzymes (like the ATPases!) are able to couple a thermodynamically unfavorable reaction with a compensating thermodynamically favorable one to yield an overall thermodynamically favorable reaction

How is energy released from ATP?

When a phosphate group is removed hydrolysis of phosphates

How could you increase the rate of a free energy reaction?

With the presence of an appropriate and active enzyme

Does a catalyst increase the rate of a chemical reaction without being permanently altered or 'used up'

Yes

Calvin Cycle

a biochemical pathway of photosynthesis in which carbon dioxide is converted into glucose using ATP

chemiosmotic coupling

a gradient of hydrogen ions across the membrane is used to drive an energy requiring process such as ATP production or the transport of a molecule across the membrane. Chemiosmotic coupling is important for ATP production in mitochondria, chloroplasts and many bacteria and archaea. ex. of oxidative phosphorylation 2-3 ATP are made for each NADH

Why is enzyme inhibition important?

a protease produced by the pancreas that is converted from the inactive proenzyme form (trypsinogen) to the active form (trypsin) in the small intestine

Citrate

a salt or ester of citric acid A compound that is an intermediate in the citric acid cycle (krebs cycle) Citrate chelates (binds) calcium ions, preventing blood clotting and thus is an effective anticoagulant

metabolic pathway

a series of biochemical reactions connected by their intermediates (the product of one enzyme catalyzed reaction is the substrate for the next enzyme catalyzed reaction) A series of chemical reactions that either builds a complex molecule or breaks down a complex molecule into simpler compounds.

Transformed cells

can grow continuously in cell culture (ex. HeLa cells) do not grow on a monolayer they have an initial loss of growth potential then immortal variant emerges -increased rate of growth -alterations in chromosomes -changes in cell surface molecules -capacity to divide indefinitely Immortal cells exceed the normal Hayflick Limit. They continue proliferating even after 40-60 doublings (no senescence; no death by apoptosis) do not typically differentiate

confluent cell culture

cells grow to touch one another, somewhat filling the culture dish

non-competitive inhibition

changes shape of the enzyme so it cannot bind

Photosystem

cluster of chlorophyll and proteins found in thylakoids

How can an enzyme be regulated?

controlling concentration covalent modifications allosteric regulation proteolytic cleavage compartmentalization and localization

how can you cause an energetically favorable reaction to 'go'

increase the temperature or add a catalyst to decrease the Ea requirement At any given temperature, a percentage of the molecules have the energy sufficient to meet the Ea requirement. (adding heat increases that percent)

competitive inhibition

interferes with the active site so that the substrate cannot bind

Oxidation

loss of electrons

Photosynthesis

process by which plants and some other organisms use light energy to convert water and carbon dioxide into oxygen and high-energy carbohydrates such as sugars and starches

feedback inhibition

process in which the product or result stops or limits the process inhibiters binding to the enzyme to stop or slow a reaction

what are two types of enzymes

protein (most enzymes) and RNA (ribosome) based

exergonic reaction

reaction that releases energy ATP hydrolysis is an exergonic reaction Exergonic is a favorable chemical reaction that has a negative Gibbs free energy . It indicates that an exergonic reaction is a spontaneous reaction since in this reaction Gibbs energy has a negative value.

covalent modification

regulation dependent upon adding/removing function groups or cleaving enzymes

amino acid job for the binding portion of the active site

responsible for the specificity of an enzyme

NADPH synthesis

series of electron carriers are used to transport electrons from chlorophyll to the coenzyme NADP+

Gluconeogenesis

the making of glucose from a noncarbohydrate source such as amino acids or glycerol pyruvate reactant to glucose product anabolic (building up) pathway

mitocondria

the powerhouse of the cell...this means that they help produce the energy needed to live -25% cytoplasm -2 membranes (anything not big can pass through)

carbon assimilation

the process by which primary producers convert atmospheric CO2 into carbohydrates during photosynthesis

K

the rate of chemical reactions

substrate

the reactant of an enzyme catalyzed reaction

enzyme kinetics

the study of the rates of enzyme-catalyzed reactions

catabolism and anabolism

two types of metabolism