Biochemistry: Vitamins & Cofactors p. 1 of 49 Many cofactors are derived from vitamins We justify...

Biochemistry: Vitamins & Cofactors p. 1 of 49

Many cofactors are derived from vitamins We justify lumping these two topics together because many cofactors are vitamins or are metabolites of vitamins.


What we’ll discuss

Tightly-bound metal ions as cofactors

Activator ions as cofactors

Cosubstrates Prosthetic groups

Vitamins Water-soluble vitaminsAscorbateCofactors

Fat-soluble vitamins


Family tree of cofactors Cofactors, coenzymes, essential ions,

cosubstrates, prosthetic groups:

Cofactors(apoenzyme + cofactor holoenzyme)

Essential ions Coenzymes

Activator ions(loosely bound)

Ions inmetalloenzymes

Prosthetic groups(tightly bound)

Cosubstrates(loosely bound)


Metal-activated enzymes

Absolute requirements for mobile ions Often require K+, Ca2+, Mg2+

Example: Kinases: Mg-ATP complex Metalloenzymes: firmly bound metal ions in active site Usually divalent or more Sometimes 1e- redox changes in metal


Coenzymes Organic moeities that enable enzymes to perform their function: they supply functionalities not available from amino acid side chains

Cosubstrates Enter reaction, get altered, leave Repeated recycling within cell or organelle

Prosthetic groups Remain bound to enzyme throughout Change during one phase of reaction, eventually get restored to starting state


Major cosubstrates Facilitate group transfers, mostly small groups

Oxidation-reduction participantsCosubstrate Source FunctionATP Transfer P,NucleotideS-adenosylMet Methyl transferUDP-glucose Glycosyl transferNAD,NADP Niacin 2-electron redoxCoenzyme A Pantothenate Acyl transferTetrahydrofolate Folate 1Carbon

transferUbiquinone Lipid-soluble e- carrier


Major prosthetic groups Transfer of larger groups

One- or two-electron redox changesProsth.gp. Source FunctionFMN, FAD Riboflavin1e- and 2e- redox transfersTPP Thiamine 2-Carbon transfers with C=OPLP PyridoxineAmino acid group transfersBiotin Biotin Carboxylation, COO- transferAdenosyl- Cobalamin Intramolec. rearrangements cobalaminMeCobal. Cobalamin Methyl-group transfersLipoamide Transfer from TPPRetinal Vitamin A VisionVitamin K Vitamin K Carboxylation of glu

residues


Adenosine triphosphate Synthesizable in liver (chapter 18)

Building block for RNA Participates in phosphoryl-group transfer in kinases

Source of other coenzymes


S-adenosylmethionine Made from methionine and adenosine Sulfonium group is highly reactive: can donate methyl groups

Reaction diagram courtesy of Eric Neeno-Eckwall, Hamline University


UDP-glucose Most common donor of glucose Formed via:Glucose-1P + UTPUDP-glucose + PPi

Reaction driven to right by PPi hydrolysis

Structure courtesy of UIC Pharmacy Program


NAD+ and NADP+

Net charge isn’t really >0 ;the + is just a reminder that the nicotinamide ring is positively charged

Most important cosubstrates in oxidation-reduction reactions in aerobic organisms

Structure courtesy of Sergio Marchesini, U. Brescia


Differences between them

The chemical difference is in the phosphorylation of the 2’ phosphate group of the ribose moiety

The functional difference is that NAD is usually associated with catabolic reactions and NADP is usually associated with anabolic reactions

Therefore often NAD+ and NADPH are reactants and NADH and NADP+ are products


How do we get back to the starting point? NADH is often oxidized back to NAD+ as part of the electron-transport chain

Imbalances can be addressed viaNAD Kinase (S.Kawai et al (2005), J.Biol.Chem. 280:39200) and NADP phosphatase


iClicker quiz: single question What would you expect to be the phosphate donor in the NAD kinase reaction?

(a) free phosphate (b) pyrophosphate (c) ATP (d) pyridoxal phosphate


Reduced forms of NAD(P)

Reduction occurs on the nicotinamide ring

Ring is no longer net-positive

Ring is still planar but the two hydrogens on the para carbon are not


FAD and FMN Flavin group based on riboflavin Alternate participants in redox reactions

Prosthetic groups: tightly but noncovalently bound to their enzymes

That protects against wasteful reoxidation of reduced forms

FADH2 is weaker reducing agent than NADH

These are capable of one-electron oxidations and reductions


FAD and FMN structures FAD has an AMP attached P to P

Structure courtesyPaisley University


Reaction diagram courtesy of Eric Neeno-Eckwall, Hamline University

FMN/FAD redox forms

Two-electron version: H+ + :H- transferred


Coenzyme A Reactive portion is free sulfhydryl at one end of the molecule

Can form thioester with acetate, etc.

Pantoate +-alanine = pantothenate

Structure courtesy ofMPB project, George Washington University

(ADP-3’P)

(Pantoate)

-alanine)

2-mercapto-ethylamine)


Thiamine Pyrophosphate

Based on thiamine, vitamin B1 Carboxylases and oxidative decarboxylases use this coenzyme

So do transketolases (move 2 carbons at a time between sugars with keto groups)

Thiazolium ring is reactive center:pKa drops from 15 in H2O to 6 in enzyme


TPP reactions

Diagram courtesy ofOklahoma State U.Biochemistry program

pyrimidine

thiazolium


Pyridoxal phosphate

PLP is prosthetic group for many amino-acid-related enzymes, particularly transaminations

Carbonyl group of PLP bound as a Schiff base (imine) to -amino group of lysine at active site

First step is always formation of external aldimine; goes through gem-diamine intermediate to internal aldimine


Biotin Rarity: vitamin is the prosthetic group

Used in reactions that transfer carboxyl groups

… and in ATP-dependent carboxylations


Biotin reactivity

Covalently bound to active-site lysines to form species called biocytin

Pyruvate carboxylase is characteristic reaction:

Diagram courtesyUniversity of Virginia Biochemistry


Tetrahydrofolate Primary donor of one-carbon units(formyl, methylene, methyl)

Supplies methyl group for thymidylate

Dihydrofolate reductase (DHFR) is an interesting drug target Methotrexate as cancer chemotherapeutic: cancer needs more thymidylate than healthy cells

Trimethoprim as antibacterial:Bacterial DHFR is somewhat different from eucaryotic DHFR because bacteria derive DHF from other sources; humans get it from folate


THF structure and function

Figure courtesy horticulture program, Purdue


Cobalamin Largest B vitamin Structure related to heme but missing one carbon in ring structure

Cobalt bound in core of ring system

Involved in enzymatic rearrangements Catabolism of odd-chain fatty acids Methylation of homocysteine Reductive dehalogenation


Adenosyl-Cobalamin

Diagram courtesy of Swiss Food News

“Missing” carbon

ReactiveCo-C bond


Lipoamide Protein-bound form of lipoic acid Contains five-membered disulfide ring

Covalently bound via amide to protein lysine sidechain

Involved in swinging arm between active sites in multienzyme complexes

Disulfides break periodically Example: pyruvate dehydrogenase complex


Lipoamide 2e- reduction Cf. Scheme 7.6: thioester starting point

Fig. Courtesy Biochem and Biophysics program, Rensselaer


iClicker revisited

Which coenzyme would you expect would be required for the reactionoxaloacetate + glutamate aspartate + -ketoglutarate?(a) ascorbate(b) PLP( c) thiamine pyrophosphate(d) NAD(e) none of the above


Vitamins: necessary micronutrients that cannot be synthesized internally What’s a vitamin for one organism is not for another

Primates and some rodents are the only vertebrates that don’t synthesize ascorbate

E.coli can make almost everything given energy and sources of atoms


Why wouldn’t organisms make everything?

Complex metabolites require energy for synthesis

Control of their synthesis is also metabolically expensive

Cheaper in the long run to derive these nutrients from diet


Vitamins: broad classifications Water-soluble vitamins

Coenzymes or coenzyme precursors Non-coenzymic metabolites

Fat-soluble vitamins Antioxidants Other lipidic vitamins

Biochemistry: Vitamins & Cofactors p. 1 of 49 Many cofactors are derived from vitamins We justify...

Documents

Transcript of Biochemistry: Vitamins & Cofactors p. 1 of 49 Many cofactors are derived from vitamins We justify...