23-24: BIOORGANIC MECHANISMS Flashcards
cofactors
-either organic molecules (coenzymes) or ions (usually metal ions) that are required by enzyme for activity
coenzyme
- low Mr organic molecule which transfers chemical groups, hydrogen or e
- often derived from a vitamin
- some come in and out of the active site as a cosubstrate (e.g. ATP)
prosthetic group
-cofactor that is bound tightly to the enzyme
apoenzyme/holoenzyzme
- apoenzyme: inactive enzyme before addition of cofactor
- holoenzyme: active enzyme and cofactor
vitamins
-small organic molecules (or set of related molecules) that are essential for growth but are unable to be synthesized by the organism so must be present in the diet
vitamins (B)
- B1: thiamine; formation of coenzyme thiamine pyrophosphate (TPP); leads to beri-beri; important catalytic centre is thiazole ring
- B2: riboflavin; formation of coenzymes FMN/FAD; leads to ariboflavinosis; important part is 3-ring system
- B3: niacin; formation of coenzymes NAD/NADP; leads to pellagra; gives rise to nicotinamide ring
- B5: pantothenic acid; formation of coenzyme A; leads to paresthesia
- B6: pyridoxine; formation of coenzyme pyridoxal phosphate (PLP)
- B7: biotin; coenzyme when covalently attached to a carrier protein
coenzymes that can be made in human body
- haem group; can exist in different forms; commonality is they all have 4 pyrrole rings ligated to central ion; substituents in the ring differ and give rise to different properties
- can make:
- lipoic acid; important in pyruvate DH complex
- Moco; involved in hydroxylation reactions because of Molybdenum ion
- ubiquinone
flavin ring system
- flexible
- undergoes 1e/1H+ reduction twice
- or straightforward H- reduction followed by protonation
Pyruvate dehydrogenase complex
- connects glycolysis to TCA cycle
- oxidative decarboxylation reaction
- E1 pyruvate dehydrogenase; E2 dihydrolipoyl transacetylase; E3 dihydrolipoyl dehydrogenase
- pyruvate comes into multi-enzyme complex; E1 active site and reacts w/TPP to allow decarboxylation to occur; 2C carbanion formed is cov. attached to TPP and is deprotonated to form intermediate hydroxyethyl-TPP
- in E2; lipoamide cofactor (important part is cyclic structure w/diS bridge; derived from lipoic acid; cov. attached to Lys on E2) in ox. form is an electrophile; carbanion Nu attack on S to reduce the diS bridge; formed acetyl-dihydrolipoamide; CoA comes in and Nu attack at carbonyl group to form acetyl-CoA (eject S as S-) and leaves behind fully reeduced lipoamide cofactor
- E3 enzyme is to reoxidise the lipoamide cofactor; 2e are abstracted to reform diS bridge and passed on to FAD and a diS bridge between 2 Cys in the enzyme becomes reduced; hydride transfer to regenerate FAD and produce NADH
why is decarboxylating pyruvate difficult?
-in a-keto-acid cannot use a-keto group as an e sink
needs cofactors:
-TPP (thyamine pyrophosphate); acts as temporary e sink (N+); key feature is thiazole ring which has acidic H that can deprotonate so under phsyiological conditions TPP can form fairly stable carbanion; can form because have delocalisation of charge onto N+ and some d-orbitals on the S; overall pKa value is about 10
importance of having a large enzyme complex
- allows for coordinated catalysis of a complex multi-step reaction
- proximity of reaction centres increases overall rate
- increases the effective concentration of reactants
- intermediates remain tethered to the complex
- minimises side reactions
substrate/metabolite channeling
- the passing of the product of one enzyme directly to another enzyme/active site without its release into solution
- can occur in stable multi-enzyme complexes such as PDH or in transient assemblies in vivo that form a metabolon
(metabolon = forms enzymes interact in a relatively weak/transient way)
mechanism of ethanol fermentation:
- pyruvate decarboxylase converts pyruvate to acetaldehyde releasing CO2 (non-ox decarb)
- decarb of a-keto-acid requires TPP cofactor
- analogous to E1 in PDH - acetaldehyde is reduced by alcohol DH using NADH to produce ethanol
- mechanism of redox regulation; recycling NAD+
- mechanism involves direct hydride transfer to the pro-R H of NADH to the re face of acetaladehyde
pyruvate carboxylase
- bicarbonate activated through phosphoryl transfer reaction involving ATP; ADP ejected to form carboxy-phosphate intermediate (more reactive); biotinyl-enzyme (carrying biotin) swings into active site; Nu attack on CO2 thats freed in close proximity to biotin to form carboxybiotinyl-enzyme (swinging-arm mechanism)
- swings out into second domain where you have pyruvate to form enolate derivative of biotin and then forms pyruvate enolate which allows Nu attack on CO2 to form oxaloacetate
