TCA Cycle

Question 1

Decarboxylation Reactions

Answer 1

• Beta keto acids: CO2 comes off 2 carbons away from ketone
  
  • Spontaneous w/o enzyme
  • Resonance stabilized
• Alpha keto acids: CO2 removed 1 carbon from the ketone
  
  • Requires enzyme b/c unstable product
  • No resonance, no e- sharing, unstable carbanion

Question 2

Pyruvate to Acetyl CoA

Answer 2

• 1st step in TCA cycle
• Mediated by Pyruvate Dehydrogenase Complex
• Many coenzymes:
  
  • Thiamine
  • Lipoate
  • FAD and NAD+
• Oxidation reaction w/ CO2 removal

Question 3

Thiamine

Answer 3

• Vitamin B1 derived
• Thiazole ring that is reactive and resonance stabilizes
  
  • Allows carbanion formation that attacks pyruvate
• Thiamine pyrophosphate: binds enzymes and interacts w/ arginines to bind thiamine:

Question 4

PDH Complex

Answer 4

• 3 enzymes:
  
  • E1: pyruvate decarboxylase (TPP)
  • E2: transacetylase (Lipoic acid)
  • E3: dihydrolipoyl dehydrogenase (FAD)
• Irreversible reaction
  
  • CO2 never build up b/c lungs remove
  • enzyme only one direction
• Regulation by phosphorylation and dephosphorylation

Question 5

PDH Complex Regulation

Answer 5

• Action by kinase and phosphatase
• Kinase phosphorylates and inactivates
  
  • Phosphorylates a serine on E1
  • Inhibited by ADP and Pyruvate
  • Activated by AcCoA & NADH
• Phosphatase dephosphorylates and activates
  
  • Activated by Ca2+

Question 6

Beri Beri Syndrome

Answer 6

• Neurological disorder caused by Thiamine deficiency
• Symptoms: lethargy, fatigue, CV, CNS, and GI issues
• Seen commonly in alcoholics and patients w/ eating disorders
  
  • Alcohol blocks thiamine uptake

Question 7

Leigh Syndrome

Answer 7

• Neurodegenerative disorder
• Onset @ 1-5 yr
• Symptoms: weakness, hypotonia, ataxia, spasticity dyspnea, opthalmoplegia, optic atrophy
• X-linked recessive
• Oxidative phosphorylation mutations causing lactic acid build up

Question 8

TCA Step 1

Answer 8

• Acetyl CoA + oxaloacetate converted to Citrate
• Mediated by citrate synthase
• ACoA comes from Acetate, pyruvate, FA, ketone bodies, AA
• H2O goes in and CoASH comes out

Question 9

TCA Step 2

Answer 9

• Citrate to Isocitrate
• Mediated by aconitase
• Moves -OH from C3 to C4

Question 10

TCA Step 3

Answer 10

• Isocitrate to Alpha-ketogluterate
• Mediated by isocitrate dehydrogenase
• Oxidation of alcohol to ketone
  
  • e- go to NAD+ to form NADH
• Decarboxylation: loss of CO2

Question 11

TCA Step 4

Answer 11

• Alpha ketogluterate to Succinyl CoA
• Mediated by alpha-KG dehydrogenase
• Similar to PDH mechanism
• Cofactors: TPP, lipoate, FAD
• Decarboxylation: loss of CO2
• CoASH enters and binds to alpha carbon

Question 12

TCA Step 5

Answer 12

• Succinyl CoA to Succinate
• Mediated by succinate thiokinase
• Thioester from SCoA cleaved to generate GTP
• Forms mixed anhydride

Question 13

TCA Step 6

Answer 13

• Succinate to Fumarate
• Mediated by Succinate dehydrogenase
• FAD converted to FADH2
• Forms double bond

Question 14

TCA Step 7

Answer 14

*Fumarate to Malate
*Mediated by Fumarase
*Addition of water to form alcohol on beta carbon
*

Question 15

TCA Step 8

Answer 15

• Malate to Oxaloacetate
• Mediated by malate dehydrogenase
• Alcohol oxidized to ketone
• NAD+ reduced to NADH

Question 16

Regulation of TCA cycle

Answer 16

1. Rate of ATP hydrolysis controls rate of ATP synthesis
   
   • controls rate of NADH oxidation in ETC
2. NAD+/NADH ratio
   * Citrate inhibits citrate synthase
   * Isocitrate DH inhibited by NADH and stimulated by ADP and Ca2+
   * alpha KG DH inhibited by NADH and stim by Ca2+
   * Malate DH inhibited by NADH

Question 17

TCA anaplerotic Reactions

Answer 17

• Forms intermediates that need regeneration
• During starvation: AAs broken to glucose (gluconeogenesis) and FA broken to form AcCoA
• AcCoA from FA stimulates breakdown of pyruvate to oxaloacetate
  
  • mediated by pyruvate carboxylase
• Fed state: lots of AcCoA and citrate w/ no cycle so citrate goes to FA