BRIDGING REACTIONSTEP 2

Fall 2013BIOT 309

TRANSITION OR BRIDGING REACTIONConnects glycolysis to citric acid/Kreb’s Cycle

OVERALL REACTION

2 pyruvate + 2 NAD+ + 2 CoA-SH (coenzyme A)

2 acetyl-CoA + 2 NADH + 2 H+ + 2 CO2

CONNECTION TO OTHER BIOLOGY: Where else is CO2 made?

TRANSITION REACTION

3 carbon

2 carbon

Co A

STEP 3 AEROBIC RESPIRATION:Krebs Cycle

BIOT 309Fall 2013

Tricarboxylic Acid Cycle = Krebs Cycle =

Citric acid Cycle

THE TCA CYCLE

• Converts acetyl CoA (from pyruvate via bridging reaction) to CO2

• Provides small amounts of energy in the form of GTP/ATP

• Collects electrons and stores as NADH and FADH2 Electron Transport Chain (ETC)

• Provides intermediates for other pathways• Occurs in cytoplasm

KREB’S CYCLESummary Reaction:

acetyl-CoA + 3NAD+ + FAD + GDP + Pi + 2H2O

——>

2CO2 + HSCoA + 3NADH + FADH2 + GTP + 2H+

TCA/KREB’S CYCLE

CITRIC ACID CYCLE

CHEMICAL REACTIONS

Key Equation: Δ G0 = -RTlnKeq

• Δ G0 = Gibb’s standard free energy change, distance from equilibrium, (expresses driving force of reaction)

• Keq =[products]/[reactants]; measurable• R= gas constant• T = absolute temperature (Kelvin)

BIOCHEMICAL REACTIONS

• Instead of Δ G0, Δ G0’ is used • Δ G0’ = standard free energy change at pH 7.0

= biochemical standard free energy

Remember: enzymes, cofactors

• Lower activation energy• Accelerate reaction• Organize and control

reaction• Recover energy in new

chemical forms and make it available for other uses

Gibb’s Free Energy

• if Δ G0’ is negative, reaction goes forward spontaneously; - products have less energy than reactants

• if Δ G0’ is ~ 0, reaction is at equilibrium• if Δ G0’ is positive, reaction does not go forward

spontaneously

• Δ G0’ of two or more reactions is calculated by adding reactions and the Δ G0’ of the reactions

• CAVEAT: Δ G0 values shown in next slides will not be true under all circumstances, could be different for prokaryotes and eukaryotes

KREB’S CYCLE, step 1

Citrate SynthaseAldol Condensation, X

2 C 4 C 6 C

KREB’S CYCLE, step 2

AconitaseDehydration, Fe-S

KREB’S CYCLE, step 3

AconitaseHydration, 4Fe-4S

Steps 2 & 3 combinedSTEPS 2 & 3 done by one enzyme

aconitaseObserve that:• Step 2: dehydration generates (double bond)

intermediate (cis-aconitate)• Step 3: dehydration moves position of OH

group

PRINCIPLE & EXAMPLE:

Δ G0’ of overall reaction is calculated by adding reactions and the Δ G0’ of the reactions*:

• Applied to 2 or more reactions, e.g., all of EMP or TCA

Δ G0’ = +2 kcal/mol

Δ G0’ = -0.5 kcal/mol

Δ G0’ = +1.5 kcal/molcitrate isocitrate

citrate cis-aconitate

cis-aconitate isocitrate

KREB’S CYCLE, step 4

Isocitrate Dehydrogenase2 step reactionOxidative decarboxylation, Mg2+ or Mn2+

NAD+

NADH, H+

6 C 5 C

SPONTANEOUS

KREB’S CYCLE, step 5

α-Ketoglutarate Dehydrogenase Complex

Oxidative Decarboxylation, TPP, Lipoic Acid, FAD

NAD+ +

CoA-SH

NADH, H+

5 C 4 C

SPONTANEOUS

KREB’S CYCLE, step 6

Succinyl CoA SynthetaseSubstrate Level Phosphorylation, FAD, TPP, Lipoic Acid

GTP converted into ATP by nucleoside diphosphate kinase

KREB’S CYCLE, step 7

Succinate DehydrogenaseOxidation, FAD & FeS

Why FAD? • alkane oxidation poorly

exergonic and can’t reduce NAD+

KREB’S CYCLE, step 8

Fumarate HydrataseHydration, Fe-S

KREB’S CYCLE, step 9

Malate DehydrogenaseOxidation

Δ G0’ = +7 kcal/mol

*

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KREB’S CYCLE !!!Summary Reaction:

acetyl-CoA + 3NAD+ + FAD + GDP + Pi + 2H2O

——>

2CO2 + HSCoA + 3NADH + FADH2 + GTP + 2H+

Transition Reaction + Kreb’s Cycle

Summary Reaction:

1 pyruvate + 4 NAD+ + 1 FAD + 1 GDP + 1 Pi

——>

4 CO2 + 4 NADH + 4 H+ + 1 FADH2 + 1 GTP(1 ATP)

EMP + TR + TCA

Summary Reaction:

GLUCOSE + 2H20 + 10 NAD+ 2 FAD +

4 ADP + 4 Pi ——>

6 CO2 + 10 NADH + 10 H+ + 4 ATP + 2FADH2

GLYOXYLATE CYCLEKREBS CYCLE ALTERNATIVE

BIOT 309Fall 2013

GLYOXYLATE SHUNT/CYCLE

• By-passes 2 decarboxylation steps in TCA making possible– net formation of succinate, oxaloacetate, and other cycle

intermediates from acetyl-CoA• Retains the two carbons lost in decarboxylation steps

with each turn of TCA• => net synthesis of oxaloacetate, a four-carbon

molecule, because each turn of the cycle incorporates two molecules of acetyl-CoA– Oxaloacetate used for other purposes

GLYOXYLATE SHUNT/CYCLE

• Allows many bacteria to metabolize two-carbon substrates such as acetate

FOR EXAMPLE:E. coli can be grown in a medium that provides

acetate as the sole carbon source. E. coli synthesize acetyl-CoA, then uses it for energy

production (via the citric acid cycle)

GLYOXYLATE SHUNT/CYCLE

• Some enzymes in common with TCA • BUT has two exclusive enzymes not in TCA

– isocitrate lyase: cleaves D-isocitrate to glyoxylate and succinate

– malate synthase: forms L-malate from glyoxylate and acetyl-CoA

GLYOXYLATE SHUNT/CYCLE

• Used when the principal or sole carbon source is a C2 compound (acetate, ethanol).

• Fat catabolism produces acetyl CoA which feeds into other catabolic reactions and produces energy