BIOCHEM EXAM: GLYCOLYSIS

2

Glycolysis generates __ pyruvate(s)

TCA Fermentation

List the two pathways pyruvate can enter once synthesized

2 4

__ ATP are consumed during glycolysis, and __ are produced

-74 kj/mol

What is the overall delta-G for glycolysis?

Hexokinase

1st enzyme of glycolysis

Hexokinase

transferase that uses a random bi-bi mechanism, delta-G’ = -16.7 kj/mol, phosphorylates glucose

phosphorylation

_______ by hexokinase adds a negative charge to keep glucose inside the cell

induced fit

Glucose and hexokinase bind via this substrate binding hypothesis

Mg2+

this cation in the hexokinase active site shields ATP’s negative charges to prevent hydrolysis

Glucokinase (HK IV)

isozyme of hexokinase found in liver, highly specific for glucose.

HK IV

this hexokinase isozyme is half the size of the others, one domain is active, the other is an allosteric regulator

phosphoglucoisomerase

2nd enzyme of glycolysis

phosphoglucoisomerase

this isomerase of glycolysis converts glucose-6-phosphate to fructose-6-phosphate

1, taking, 2, gives, 1, 1, 2, aldehyde, ketone

phosphoglucoisomerase uses H388 and K518 to attack the C_ bond, ____ the ring of glucose while ____ a proton from C_, then the enzyme ____ a proton to C_, moving the carbonyl from C_ to C_, from a _____ to a _____

Neuroleukin

Phosphoglucoisomerase secreted by T cells that promotes spinal/sensory neuron survival

Autocrine motility factor

Phosphoglucoisomerase secreted by tumor cells that stimulates cancer cell migration

Differentiation and Maturation Mediator

Phosphoglucoisomerase secreted by T cells that promotes leukemia cell differentiation

Homodimer

phosphoglucoisomerases structure can be described as a:

Phosphofructokinase

3rd enzyme of glycolysis

phosphofructokinase

This enzyme of glycolysis is highly regulated, as it represents a commitment to the pathway

phosphofructokinase

This glycolytic transferase converts a-fructose-6-phosphate (F6P) → a-fructose-1, 6-bisphosphate (FBP)

pyruvate

Phosphoglycerate kinase functions similarly to _______ kinase

phosphoglycerate kinase

pyruvate kinase functions similarly to _______ kinase

ATP

____ is an allosteric inhibitor of phosphofructokinase (PFK)

decrease

When ATP is increased, PFK activity should ______

Activator increases

F26BP is an ________ of phosphofructokinase, the rxn velocity _____ when more F6P substrate is added

F26BP

NOT a metabolite of glycolysis, made by PFK-2, hormonally regulated activator of glycolytic PFK

homotetramer 4 4

PFK structure can be described as a __________ with __ catalytic sites and __ known allosteric sites

phosphofructokinase (PFK)

this glycolytic enzyme is regulated by citrate, F26BP, AMP, ADP, and others, affecting not just glycolysis

T state

This is the inhibited form of phosphofructokinase

R state

This is the activated form of phosphofructokinase

fructose bisphosphate aldolase (FBA)

4th enzyme of glycolysis

fructose bisphosphate aldolase (FBA)

this glycolytic lyase converts linear F-1,6-BP to dihydroxyacetone phosphate (DHAP) and alpha-glyceraldehyde-3-phosphate (G3P)

Class 1

this class of fructose bisphosphate aldolase is found in animals, and uses covalent Schiff base chemistry

Class 2

this class of fructose bisphosphate aldolase is found in bacteria and fungi, and uses metal ion catalysis (usually Zn²⁺)

cyanobacteria

example of an organism that can make both classes of fructose bisphosphate aldolase:

Schiff Base

a covalent imine (N=R) linkage between a sugar and an enzyme

Lysine carbonyl 2 schiff base

______ of the FBA active site attacks the ______ of C_ of F16BP, forming a _____ _____

aspartate proton 4 3 G3P enamine

an active site base of FBA, like _______, removes a _____ from C_ of F16BP, triggering the cleavage of the bond between that carbon and C_, resulting in relase of ___ as a free molecule, and leaving a covalently linked _____ attached to the enzyme

enamine imine

after release of G3P, the ________ still attached to the enzyme is tautomerized into a(n) ________

water dhap

_____ attacks the schiff base of FBA, hydrolyzing it, releasing ____

bind schiff base asp G3P enamine imine water schiff base dhap

Steps of FBA mechanism: ____ → form ____ ____ → ___ cleaves C3-C4 bond → ___ released → ______ intermediate to ______ intermediate → _____ hydrolyzes the ____ ____ → ____ released

2 Zn2+ 2 schiff base 1

Class __ FBAs use a ___ cation to polarize the carbonyl on C_, rather than the covalent ____ ___ formed in Class _ aldolases

Triose Phosphate Isomerase (TPI)

5th enzyme of glycolysis

Triose phosphate isomerase (TPI)

this glycolytic isomerase converts DHAP to G3P

DHAP G3P

Triose phosphate isomerase (TPI) converts _____ → _____

glu165

TPI active site base that takes a proton from C1 of DHAP

DHAP enediol G3P

TPI Mechanism: ____ →/← ________ intermediate →/← ____

Glu165 1 DHAP enediol 1 carbonyl 2 1 G3P

in TPI mechanism: ____ takes a proton from C_ of ___, forming an _____ intermediate (C=C with 2 OH groups), then a base of the active site takes a proton from the C_’s hydroxyl group, which forms a _______ and returns the glutamine’s proton to C_ instead of C_, forming ____

2

How many moles of ATP are consumed for each mole of glucose in the energy investment phase of glycolysis?

0

How many moles of ATP are produced for each mole of glucose in the energy investment phase of glycolysis?

2

How many moles of G3P are produced for each mole of glucose in the energy investment phase of glycolysis?

G3P dehydrogenase

6th enzyme of glycolysis

G3P Dehydrogenase

This glycolytic oxidoreductase is constitutively expressed in most tissues and cells, used as a positive control for gene expression studies, converts G3P → 1,3-bisphosphoglycerate (BPG)

G3P 1,3-bisphosphoglycerate (BPG)

GA3PDH converts _____ → ______ via hydrolysis

NAD+ NADH

Reduction of ____ to ____ is a way of moving electrons around the cell

NADH ETC 1

Electrons from ____ eventually enter the ___ complex __

G3P Cys Aldehyde 1 His 1 Cys thiohemiacetal his oxidizes 1 hydride NAD+ thioester 2 NADH phosphate 1,3-BPG G3P

GA3PDH mechanism: ___ enters, ___ residue attacks the ______ carbon (C_), reducing the aldehyde, and deprotonating the ___ residue, bonding C_ to the ___ residue, and creating a ___________ intermediate. The ___ residue then re-_________ C_, regenerating the aldehyde, and losing a _______ to ____, creating a ______ linkage between the enzyme and the _ carbon remainder of G3P. _____ is released, then inorganic ________ attacks, and the enzyme releases _______, and the enzyme is ready for more ___ to enter.

phosphoglycerate kinase (PK)

7th enzyme of glycolysis

phosphoglycerate kinase (PK)

This enzyme’s reaction is an example of substrate level phosphorylation

substrate level phosphorylation

a method of ATP synthesis by directly transferring the phosphate from a high energy substrate to ADP

phosphoglycerate kinase (PK)

This is the first glycolytic enzyme to produce ATP

phosphoglycerate kinase (PK)

this glycolytic transferase

phosphoglycerate kinase (PK)

this glycolytic transferase converts 1,3-bisphosphoglycerate (BPG) → 3-phosphoglycerate (3-PG), transferring a phosphate to ADP to produce ATP

Phosphoglycerate Mutase (PGM)

8th enzyme of glycolysis

Phosphoglycerate Mutase (PGM)

this glycolytic isomerase converts 3-Phosphoglycerate (3-PG) → 2-Phosphoglycerate (2-PG)

3-PG phosphor-histidine phosphate 2 3-PG 2,3-PG His 3 2-PG

PGM Mechanism: ___ binds, active site ______-_______ donates its _______ to C_ of ___, generating the high energy ___________ intermediate, dephosphorylating the ___ residue, which then attacks the phosphate of C_, re-phosphorylating it for the next substrate, and releasing the newly formed _______

His phosphate 2 phosphate 3

PGM Mechanism can be simplified as: ___ residue gives a _______ to C_, and then takes the ________ from C_

Enolase

9th enzyme of glycolysis

Enolase

this glycolytic lyase converts 2-Phosphoglycerate (2-PG) → Phosphoenolpyruvate (PEP) via a dehydration reaction, removing the alcohol from the terminal carbon, and forming a high-energy phosphate compound

Lys deprotonate 2 negative mg2+ glu 3 hydroxyl water 2 3 PEP

Enolase mechanism: ___ residue acts as general base to ________ C_, forming a _______ charge stabilized by ____ ions. ___ residue donates a proton to C_’s _______ group, releasing ______, generating the C_←>C_ double bond, and releasing ___

Pyruvate Kinase

10th enzyme of glycolysis

Pyruvate Kinase

this glycolytic transferase converts Phosphoenolpyruvate → pyruvate, transferring a phosphate to ADP to generate ATP

PEP oxygen phosphate PEP ATP pyruvate

Pyruvate Kinase Mechanism: ___ and ADP bind, _______ of ADP attacks the _________ of ___, yielding ___ and ______

HK PFK PGK PK

Name all the glycolytic phosphotransferases:

ATP Glucose

Hexokinase transfers a phosphate from ______ to ______

ATP Fructose-6-Phosphate

Phosphofructokinase transfers a phosphate from ______ to ______

1,3-Bisphosphate ADP

Phosphoglycerate Kinase transfers the 1-phosphate of ______ to ______

phosphoenolpyruvate ADP

Pyruvate Kinase transfers a phosphate from ______ to ______

deprotonation base

all glycolytic isomerase reactions start with the _________ of a substrate by a general _____

PGI TIM

These 2 glycolytic isomerases use both a general base and acid in their active sites

PGM

this glycolytic isomerase rearranges a phosphate linkage using a phosphohistidine amino acid

PGI TIM

Both of these glycolytic isomerases create an enediol intermediate

PGM

This glycolytic isomerase creates a 2-3-bisphosphoglycerate intermediate

PGM

The reaction of this glycolytic isomerase closely resembles that of phosphotransferase chemistry

Aldolase

This glycolytic lyase forms a Schiff base between enzyme and substrate, cleaving a C-C bond

GAPDH

this glycolytic oxidoreductase forms a thiohemiacetal with a cysteine residue, reducing NAD+ → NADH

Enolase

this glycolytic lyase deprotonates at the alpha-carbon (C-H bond), forming an enolate intermediate, resulting in C-O bond cleavage

Transferase

Hexokinase is a:

Isomerase

Phosphoglucoisomerase is a:

Transferase

Phosphofructokinase is a:

Lyase

Aldolase is a:

Isomerase

Triose Phosphate Isomerase is a:

Oxidoreductase

Glyceraldehyde-3-Phosphate Dehydrogenase is a:

Transferase

Phosphoglycerate Kinase is a:

Isomerase

Phosphoglycerate mutase is a:

Lyase

Enolase is a:

Transferase

Pyruvate Kinase is a: