EXAM 2 Flashcards
Glycolosis
catabolic pathway universal to virtually all organizsm. involves a series of enzyme catalyzed reactions within the CYTOSOL which phosphorylate, modify and split 6-carbon glucose molecule to eventually produce 2 molecules of 3- carbon pyruvate. ATP is consummed but overall gain is 2 ATP Key pathway for metabolsim of carbohydrates
how does glucose go from blood to cell?
via specific glucose transporter protein GLUT1. this is a pssive glucose transporter that does not involve a coupled mouvement of any other ion or molecule
how does glucose transport differ in the gut?
in gut epithelial cells the transport is an active mech coupled with the co-transport of Na+ using the existing Na-gradient to drive glucose from the gut lumen into the cell lumen against the concentration gradient of glucose
Where does glycolosis occur?
cytosol of all tissues
what happens to glucose once it passively enters the cell?
it is rapidly phophoylated at the 6-th position by HEXOKINASE( or glucokinase)
what 4 pathways can the phosphorylated glucose enter?
1-glycolytic pathway( oxidation to provide energy-leads to formation of pyrvate)
2-pentose phosphate pathway( Oxidation, to produce cytosolic reducing power for synthesis, nucleotides or energy)
3-glycogen synthesis pathway(to store glucose for utilization at another time)
4-Synthesis of complex polysaccharides(cell matrix and cell wall polysaccharides)
why is glucose phosphorylatd
1- to trap glucose within the cell(hence permit more glucose to passively diffuse in)
2-conserve energy:the phosphate that is donated to glucose from ATP will be used to regenerate ATP
3-“Tagging” of glucose all the glycolytic pathway intermediates are phosphorylated; the enzymes of the pathway recognize only these phosphorylation derivatives
what enzymes phosphorylate glucose?
Hexokinase- most tissues, non specific, low Km(0.1mM) product inhibited( Glucose-6-phosphate)
2- GlucoKinase:only liver, glucose specific. high Km(10mM) no product inhibition(not saturated nor does build-up of its product reduce its activity there for it can continue to phosphorylate the excess glucose and remove it from the blood)
what are the 2 phases of glycolosis?
the energy investement phase( cleavage of hexose chain) and the energy pay off stage
steps of phase 1 glycolysis?
reaction 1: phosphorylation of glucose =the first ATP INVESTEMENT (hexokinase)
Reaction 2: Glucose-6-phosphate conversion to fructose-6-phosphate=isomerization of glycose-6-phosphate (phophohexose isomerase)
Reaction 3: fructose 6 phosphate phosphorylation to fructose 1,6-biphosphate= the second ATP INVESTEMENT (phophofructokinase)
Reaction 4: cleavage of fructose 1,6-biphosphate=production of 2 triose phosphates( aldolase)
Reaction 5: triose phophate interconverstion= isomerization of dihydroxyacetone phosphate ( triose -phosphate isomerase)
what is the outcome of the 1st phase of glycolysis?
energy is invested in the form of 2 ATP per molecule of glucose the 6 carbon glucose molecule splits into two 3-carbon molecules of glceraldehyde 3 phosphate
what occurs in the pay off phase of glycolysis?
Oxidative conversion of glyceraldehyde 3- phosphate to pyruvate and couplded formation of ATP and NADH
what are the irreversable steps during the preparatory phase of glycolysis?
hexokinase reaction and the phosphofructokinase reaction. the PFK reaction is the committing step to glycolysis
steps of phase 2 of glycolysis
Reaction 6: Glyceraldehyde 3 phosphate oxidation to 1,3-bisphosphoglycerate= generation of the first energy rich compound (glyceraldehyde 3 P dehydrogenase)
Reaction 7: phophate transfer from 1,3 bis phosphoglycerate to ADP= the first substrate level phosphorylation to PRODUCE ATP(phosphoglycerate kinase)
Reaction 8: 3- phosphoglycerate to 2- phosphoglycerate conversion=preparation for syntehesis of the next energy-rich compound(phophoglycerate mutase)
Reaction 9: Dehydration of 2-phophoglycerate conversion to phosphophenalpyruvate=synthesis of the second high-energy compound (Enolase)
Reaction 10: phosphoryl transfer from phosphoenol pyruivate to ADP= the second substrate level phosphorylation to produce ATP (Pyruvate kinase)
how many molecules of ATP are formed from glycolysis
4 per molecule of glucose and 2 per molecule of glyceraldehyde-3-phosphate
is the pay off phase of glycolysis reversible?
yes these reactions are substrate limited and can be easily reversed
what enzyme catalyzes reaction 10 in glycolysis? is this reaction in equilibrium?
pyruvate kinase. reaction is far from equilibrium in cells and has a high driving force in the forward direction
what is the significance of the energy levels of PEP hydrolysis?
it is almos 2 that for hydrolysis of the terminal phosphate from ATP, PEP has the highest phosphate transfer potential of any biological compund hance the energy from its hydrolysis is sufficient to easily drive synthesis of ATP from ADP with plenty of energy lef over to provide a substantial driving force in the forward direction
why does PEP have such a high energy of hydrolysis?
because the initial product of PEP hydrolysis is enol-pyruvate which is very unstable and spontanesously froms the Keto-form
what is the other high energy phosphate formed during the pay off phase of glycolysis at reaction 6?
glyceraldehyde 1, 3 biphosphate. this is an energy conserving reaction in which the energy of oxidation of an aldehyde to an acid is conserved by forming a high energy acyl-phosphate (50% higher energy provided by the hydrolysis of this compound than by ATP)
what are the reactions in glycolysis that do NOT operate at equilibrium?
HK, PFK and PK
what is the net production of glycolysis ?
2 ATP 2 NADH during the conversion of 1 molecule of glucose into 2 molecules of pyruvate
what are the two metabolic fates of pyruvate?
1- entry into the citric acid cycle(via pyruvate dehydrogenase)
2-anaerobic reduction to lactate and the regeneration of NAD+( via lactate dehydrogenase)
what is the fate of pyruvate in muscles cells during rigorous excercise ?
MADH from reaction 6 of glycolysis cannot be converted back. lack of NAD+ inhibits the reaction. so pyruvate will be converted into lactate(recution step coupled to the oxidation of NADH back to NAD+–> glycolisis can continue on and produce ATP without any net oxidation or reduction
what is ATP’s effect on phophofructokinase in glysolysis?
ATP is an important negative allosteric regulator of the enzyme. high levels of ATP considerably increase the Km of the enzyme–> increase ATP in cell, decrease in glycolysis
what positively regulates activity of PFK?
ADP, AMP and fuctose 2,6 bisphosphate
what other carbohydrates can enter into glycolysis
fructose (through hexokinase)
Galactose (attached to UDP then epimerized to glucose)
Mannose *ophosphorylated by hexokinase then changed to fuctose-6-p by the action of phosphomannose isomerase)
what is the role of the Pentose Phosphate Pathway(PPP)
provides an alternative route to oxidation of glucose-6-phosphate
pahway yield NADPH which is usually used for reductive biosynthesis
NADPH protects agains oxidative damage due to O2 and H2O2
NADPH is used to recude oxidized glutathione(GSS)
what are the products of the Pentose Phosphate Pathway (PPP)
2 NADPH and ribose-6-phophate thei is called the oxidative phase
inthe non-oxidative phase-ribose-6-phosphate can be used for nucleotide syntehsis or recycled to glucose-6 phosphate(can go into glycolysis or PPP divison regulated by levels of NADPH in the cell)
how much glucose does the body need perday?
160 grams- of that 120 grams go to the brain
what is gluconeogenesis?
when glucose is depleted in fasting times there is the generation of glucose from non-sugar carbon compounds
which amino acids cannot provide carbon for glucose synthesis?
leucine and lysine
what are important glucose precursors for gluconeogenesis?
lactate pryuvate glyceral and amino acids
how are amino acids used for net glucose synthesis?
they are catabolized to pyruvate or intermediates of the citric acid cycle that can undrego oxidation to OXALOACETATE. bothy pyruvate and oxaloacetate are readily funneled into gluconeogenesis
where does glugoneogenesis take place?
mainly in the lliver some in renal cortex too and small intesting epithelial cells`
What happens to lactate produced in active muscle cells?
the lactate is transported to the liver and converted to glucose via gluconeogenesis then glucose is realeased to the blood and returns to the muscle where it is converted into glycogen (CORI CYCLE)
what occurs in gluconeogenesis?
pyruvate is converted into glucose
are glycolysis and gluconeogenesis the same but in opposite directions?
no, they do not have identical pathways. 7 of the 10 reactions of glycolysis which have very low free energy change and can be easily reversed and therefore are shared by both pathways
what are the 3 reactions in glycolyisis that are characterized by a large negative Delta G?
1- conversion of glucose to glucose-6-phosphate to fructose1,6-bisphosphate
2-phosphorylation of fructose-6-phosphate to fructose 1,6-biphosphate by phosphofructokinase-1
3-conversion of phosphoenol-pyruvate to pyruvate by pyruvate kinase
how does gluconeogenesis deal with the 3 irreversible reactions of glycolysis?
they are bypassed by a seperate sets of enzymes
what is the first reaction that is bypassed in gluconeogeneosis?
converstion of pyruvate into phosphoenolpyruvate(PEP) there are 2 alternative pathways from pyruvate to PEP, the path depends on the glycogenic precursor : pyruvate or lactate
what is the pathway in the first bypass when pyruvate is the starting material?
it is first transported from the cytososl into the mitochondria or is generated from alanine within the mitochondria by transamination.
pyruvate then is CARBOXYLATED to OXALOACETATE(4 carbons) at the expense of an ATP.
oxaloacetate is then REDUCED to MALATEmy (mitochondrial malate dhydrogenase) at the expense of NADH
MALATE exits the mitochondrion through a specific transporter in the inner mitochonrdial memebrane
in the cytosol it is OXIDIZED back to OXALOACETATE with production of cytosolic NADH(cytosolic malate dehydrogenase)
finally OXALOACETATE is DECARBOXYLATED to PEP by phosphoenolpyruvate carboxykinase ( this is an MG2+ dependat reaction and requires GTP as a phosphoryl group donor)
what does the dexacroxylation of oxaloacetate to PEP require?
phophoenolpyruvate carboxykinase. this is an Mg2+ dependent reaction and require GTP as the phosphoryl group donor
what is the signification of the oxidisation of malate back to oxaloacetate in the cytosol?
leads to the production of cytosolic NADH from NAD+. as a resullt NADH “shuttled out” from mitochondrion without crossing the memebrane to be used later in gluconeogenesis
what is the overall reaction of the bypass of pyruvate kinase starting with pyruvate?
Pyruvate+ATP+GTP+HCO3—>PEP+ADP+Pi+CO2 (Delta G=0.9kj/mol)
how is the first bypass reaction from pyruvate irreversible?
it is actually very negative because PEP concentration is low due to its consumption in other reactikon. so this reaction is effectively irreversible in the cell
what is the bypass reaction of pyruvate kinase when lactate is the starting material?
OXIDATION of lactate to pyruvate in the cytosol of hepatocyte by lactate dehyrogenase is accompanied by the production of NADH.
this reaction provides sufficient amount ot cytosolic NADH for gluconeogenesis so there is no need to”shuttle” NADH from the mitochonrdia
after produced pyruvate is transported to the mitochonrdia it is CARBOXYLATED to OXALOACETATE pyr pyruvate carboxylase at the expense of ATP
this OXALOACETATE is DECARBOXYLATED to PEP inside the mitochondria by mitochondrial PEP carboxykinase and PEP is transported from the mitochondria into the cytosol to continue on the gluconeogenic path
where is lactate produced?
erythrocytes or anaerobic muscle
how is the reaction of phosphofructokinase-1 bypassed in gluconeogenesis?
the second reaction that cannot be utilized in gluconeogenesis is the PHOSPHORYLATION of fructose-6-phosphate by phosphofructokinase-1 PFK 1
it is bypassed by the mg2+ dependent fructose 1,6-bisphosphatase which promotes the essentially irreversible hydrolysis of the C-1 phosphate
these 2 opposite reactions are catalyzed by different enzymes and are entirely different
Fructose1,6-bisphosphate_+H2O–> Fructose6-phospphate+pi (-16.3 kJ/mol)
what is the 3rd by pass and how is it done?
the 3rd irreverisble reaction is the bypass of hexokinase
the bypass reaction , the DEPHOSPHORYLATION of glucose 6-phosphate to yield glucose is catalyzed by glucose6-phosphatase
glucose 6-phosphatate+H2O–>glucose+Pi
the Mg2+ dependant enzyme is located on the inside surface of the ER of hepatocytes and renal cells (muscle and brain tissue dont have this enzyme there for they are dependnt on the outside supply of glucose delivered through the bloodstream)
what is the overall reaction of gluconeogenesis?
2 pyruvate+4 ATP+2 GTP+2NADH+2 H+ +4 H2O–> glucose+ 4 ADP+ 2 GDP + 2 NAD+ +6 Pi
how do cells assure the irreversibility of glyconeogenesis?
this is an expensive process and much of the high energy cost is necessary to ensure the irreversibility of gluconeogenesis
what is the first control point in gluconeogenesis?
this first congtrol point determines the fate of pyruvate in the mitochondria
Acetyl-CoA is a positive allosteric moedulator of pyruvate carboxilase and a negative modulaor of pyruvate dehydrogenase
accumulation of Acetyl-CoA means the cells needs are met- it inhibits pyruvate dehydrogenase, slowing formation of acetyl-coA from pyruvate and sitmulates guconeogenesis by activating pyruvate carbozylase
what is the second control point of gluconeogenesis?
irreversible conversion of furcotse 1,6 bisphophate to fuctose 6-phosphate catalyzed by the fructose 1,6-bisphosphatase-1(FBP-1) the corresponding glycolytic reaction is catalyzed by a diff enzymes phosphofuctokinase-1(PFK-1)
simultaneous operation of both enzyme would consume ATP wihtout accomplishing any chemical or byological works, futile cycle.
to avoid this they are regulated in a coordinated and reciprocal manner allowing only one proces to be active at a time
AMP inhibits FBP ase-1 and stimulates PFK-1
PFK-1 is also stimulated by ADP but inhibited by citrate and ATP
all changes regulated by rapid allosteric mech
What are the regulatory processes triggere from outside the cell for glycolisis and gluconeogenesis?
by hormones insulin and glucagon
the hormonal regulation of both pathways is mediated by FRUCTOSE 2,6- BISPHOSPHATE (F26BP). its in allosteric mediator that stimulate PFK-1 and inhibitse FBPase-1–> stimulates glycolysis and slowing gluconeogenesis
how are levels of Fructose 2,6-bisphosphate (F26BP) regulated?
it is formed by the phosphorylation of furctose 6-phosphate catalyzed by phosphofructokinase -2 (PFK-2 and is borken down by fructose 2,6 bisphosphatase-2 ( FBPase-2) they are regulated in a reciprocal fashion by insulin and glucagon
what effect does glucagon have?
glucagon stimulates liver adenylate cyclase to sinythesie cyclic AMP that acitvates CAMP dependant protein kinase phosphorylation of PFK-2/FBPase-2 protin this phosphorylation increases FBPase-2 activity and inhibits PFK-2 activity –> lowers F26BP levels and inhibits glycolysis and simulates gluconeogenesis
–> glucagon promotes release of glucose in blood
what is the effect of insulin in regulateing glucose levels?
insuline simulates the dephosphorylation of PFK-2/FBPase-2 by a phosphoprotein phosphatase.
–> inhibition of FBPase-2 and activation of PFK-2 that increases level of F26BP–>stimulates glycolysis
how does insulin regulate transcription?
insuline acts through its membrane receptor to turn on the signaling pathway resulting in the activation of prtoein kinase B(PKB)
in response to insulin the transcription factor FOX01 –> stimulates the synthesis of gluconeogenic enzymes PEP carbizykinase and Glucose 6-phosphatase
FOX01 leaves the nucelues and is phosphorylated by activated PKB then tagged by ubiquitin
how does glucagoneffect FOX01 activity?
glucagon prevents the phosphorylation of FOX01 by the activated PKB. FOX01 remains aftive in the nucleius
what is the first step of cellular respiration?
organic fuel molecules are oxidized to yield 2 carbon fragments in the form of acetyl group of acetyl group of acetyl-coenzyme A(Acetyl- coA)
what is the second step of cellular resipiration?
the activated acetyl groups( acetyl-CoA) are fed into the citric acid cycle which enzymatically oxidizes them to CO2 the energy released is conserved in the form of reduced electron carriers NADH and FADH2
what is the 3rd step of cellular respiration?
NADH and FADH2 are oxidized and the electrons they carry are transferred through the respiratory (electron-transfer) chain to )2 the final electron acceptor ruducing it to H2O. this electron flow drives the production of energy which is conersved in the form of ATP by a process called oxidative phosphorylation
what is the significance of the citric acid cycle?
the citric acid cycle is the final common pathway for the oxidation of fuel molecules - carbohydrates, fatty acids, and some amino acids (a hub metabolsim)
how does pyruvate enter the citric acid cycle?
pyruvate is converted into Acetyl- CoA by pyruvate dehydrogenase PDH complex. this complex catalyzes irreversible oxidative decarbozylation of pyruvate resulting in the remouval of its carboxyl group in the form of Co2 and formation of acetyl-CoA. this reactikon requires the sequentia action of three diferent enzymes E1 E2 and E 3 and 5 diff coenzymes
what are the enzymes of the PDH complex?
pyruvate dehydrogenase(E1) dihydrolipoly transacetase(E 2) Dihydrolipoyl dehydrogenase(E 3) each are present in multiple copies in the complex number of copies varies between species eukaryotes ( 30 E1 and 12 E 3)
what are the co-enzymes in the PDH complex?
coenzyme A(CoA-SH)- reversibly associating coenzyme containing a reactive thiol-SH group that froms a thioester with acetate in acetyl-CoA
Nicotinamide Adenin Dinucleotide(NAD+): also a reversibly associating coenzyme it serves as an electron carrier
Thiamine Pyrophosphate(TPP) attached to E1 and serves as a catalytic cofactor
Lipoate: Attached to E2 through Lys of E2 and has 2 thiol groups that can undergo reversible oxidation to a disulfide bond it can serve both as an electron carrier and as an acyl carrier
Flavin Adenine Dinucleotide(FAD): attached to E3 it serves as an electron carrier
what is the reaction mechanism of the PDH complex?
step 1: pyruvate reacts with the bound TPP of pyruvate dehydrogenase(E1) undergoing decarboxylation to the hydroxyethyl derivative(2 carbons)
step 2: E1 trasnfers the acetyl group and 2 electrons from TPP to the oxidized form of the lipoyllysine of the core enzyme,dihydrolipyl transacetylase(E2)
step 3: in the reaction of transesterification the -SH group of CoA replaces the- SH group of E2 to yield acetyl-CoA and fully reduced form of the lipoyl group
Step 4: Dihydrilipoyl dehydrogenase(E3) promoes the trasnfer of 2 hydorgen atoms from the reduced lipoyl groups of E2 to the FAD porstetic group of E 3, restoring the eoxidized from of the lipoyllysyl group of E2
Step5: the reduced FADH2 of E3 transfers a hydride ion to NAD+ resulting in NADH
explain the “swinging arm” term?
the lipoamide of E2 provides the swinging arm that transfers the product of E1(the hydroxyethyl group) to the active site of E2 where it is oxidized to the acetyl group and trasnfered to CoA-SH
the swinging arm ins its reduced state then interacts with E 3 in order to regenerate its oxidized head group
why is the PDH complex an example of the tethering mechanism for the channeling of a substate between active sites?
1- diffusion of substrates and products between the enzymes is no required (increase in reaction speed)
2- the intermediates never leave the complex(increase in concentration and reaction speed)
3-other competing compounds are prevented from entering active sites( increase reaction fidelity)
where does the citric acid cycle take place?
mitochondria
8 steps in which the 2 carbon acetyl CoA is completely eoxidized to CO2 with most of the energy conserved in the form of NADH and FADH2
what is the 1st step of krebs cycle?
formation of citrate:
the reaction is catalyzed by citrate synthase
involved aldole condensation of acetyl-CoA with oxaloacetate to form citroyl-CoA which is then hydrolyzed to citrate and CoA
the hydrolysis of Citroyl-CoA pulls the overall reaction in the direction of citrate synthesis
the large negative Delta G is essential to the operation of the cycle due to the very low level of oxalacetate in the mitochondria
what is the 3rd step of the krebs cycle?
Oxidation of isocitrate to alpha Ketoglutarate and CO2:
the reaction is catalyzed by isocitrate dehydrogenase
this is the first OXIDATION reaction of the cycle it involved the transfer of electrons to NAD+ it is also a decarboxylation step with CO2 being released
there are 2 diff forms of isocytrate dehydrogenase one reuiring NAD+ as an electron acceptor occurs in the mitochondria and serves in the citric acid cycle the other can recognize NADP+ and is mainly used to make NADPH for reductive biosynthesis in the cytosol
large Negative delta G-imp REGULATORY STEP, rate of formation of alpha-ketoglutarate is imp in determining the overall rate of the cycle
what is the 2nd step of the krebs cycle?
Formation of isocitrate via cis-aconitate:
the reaction is catalysed by ACONITASE
the reaction involves remouval and addition of water dehydration-hydration
aconitase can promote the reversible additon of H2O to the double bond of cis-aconitate in 2 diff ways. one leading to citrate and the other to isocitrate however the reaction is pulled to the right beacuse the isocitrate is rapidly consumed in the next step of the cycle
large positive delta G but close to equilibrium in the cell because the product is kept at low concentration
what is the 4th step of the krebs cycle?
Oxidation of Alpha-Ketoglutarate to Succinyl-CoA and CO2:
the reaction is catalyzed by Alpha-Ketoglutarate dehydrogenase complex which is very semilar to the pyruvate dehhydrogenase compled. the mech is also very similar to that of the conversion of pyruvate to acetyl-CoA
this is the second coidation/dexarboxylation reactions of the cycle
NAD+serves as an electron acceptor and COA-SH is a carrier of the succinyl group
the reaction has a large neg G, REGULATED step
what is the 5th step of the krebs cycle?
Conversion of Succinyl-CoA to Succinate:
the reaction is catalyzed by Succinyl-CoA synthase or Succinic Thiokinase
this is the ONLY phosphorylation steop in the cycle that directly yields a high energy phosphjate bond. all other ATPs are gnerated indiretly through electron carriers and oxidative phosphorylation
GDP is phosphorylated to GTP , GTP is sued as a phosphoryl donor in protein synthesis and signal transduction or can easly be converted to ATP by nucleotide diphosphokinase
low Delta G-REVERSIBLE REACTION
