Lecture 3- Enzume Catalysis, Inhibition, and Regulation

how much do enzymes typically increase rxn rates?

10^6-10^14

Is delta G changed by enzymes?

No

Is eq constant changed by enzymes?

no

What can enzymes allow for

rxn to occur in mild conditions

What is considered mild conditions

37ºC and neutral PH

Are enzymes specific or general

highly specific

Can enzymes be regualted

yes

What does enzyme do to activation energy

decrease it

what is the substrate for hexokinase

glucose, not galactose

Is ethanol or methanol/ isopropanol reduced quicker by alcohol dehydrogenase

ethanol bc higher affinity

Are CYP450 broad or specific substrate

broad

How are substrates recognized at binding site

by amino acids through hydrogen bonds, ionic interactions, and hydrophobic interactions

what do enzymes do upon substate binding and why?

change 3d shape, increases reaction occurance

Example of induced fit

conformational yeast hexokinase occurs when glucose is bound, causes ATP binding and excluding of water

what cofactor is used to reduce methanol to acetone

NAD

Oxidoreductase

catalyse oxidation reduction rxn

examples of an oxidoreductase

alcohol dehydrogenase

What will you see with oxidoreductase?

NAD+ to NADH

FADH to FADH2

What do transferases do

transfer functional groups

example of transferase

kinases

what is something you might see with transferase

ATP to ADP in reactants

Hydrolase function

carry out hydrolysise

example of hydrolase

chymotrypsin (a serine protease)

What will you see with hydrolases

H2O going in + split

lyases function

cleave or form bonds (without redox), add to double bond

• forms two products

example of lyases

aldolase

Isomerases function

isomerization

example of isomerase

triose phosphate isomerase

ligases function

forms two bonds between molecules by breaking a high energy bond

• C-C

• C-O

• C-S

• C-N

what will be seen with ligase

ATP to ADP

example of ligase

pyruvate carboxylase, makes pyruvate into oxaloacetate

What does aldolase do

makes fructose 1,6-bisphosphaye into GAP and DHAP

What are the strategies for enyzyme catalysis

acid base

covalent

transition state binding

catalysis by proximity

catalysis employing cofactors

does an enzyme have to use one mechanism for catalysis

no can use multiple

acid base catalysis

protein functional group donates or accepts a proton during rxn

covalent catalysis

a reduction is accelerated through formation of covalent enzyme substrate bond

transition state binding

preferential binding of the transition state of rxn lowers EA of rxn

Catalysis by proximity

enzyme places substrates in orientation for the rxn to ocur and restricts unfavorable orientations

Catalysis employing cofactors

cofactors expand the chemical versatility of enzymes

Examples of cofactors

coenzymes, metal ions

What are the serin proteases

trypsin, elastase, chymotrypsin, thrombin

what is thrombine

serine protease involved with coag cascafe

digestive serine protease

trypsin and elastase

What does chymotrypsin make use of for peptide hydrolysis?

acid base catalysis

covalent catalysis

and transition state binding

chrymotrypsin prefers

aromatic, large bulky hydrophobic side chains like phenylalanine F, tryptophan W, tyrosine Y

Trypsin prefers

lys and arg LARGE POSIRIVE RESIDUES

elastase prefers

Ala Gly Val SMALL HYDROPHOBIC

What is formed by serine protease mechanism

oxyanion hole which stablizes the transition state

What amino acid can be present in acid base catalysis

histidine

The bacterial enzyme DNA gyrase is the target

of fluoroquinoline antibacterial drugs such as ciprofloxacin.

In the process of acting on its substrate, DNA gyrase forms

a covalent intermediate through nucleophilic attack on the

DNA backbone. Which amino acid found in DNA gyrase

forms this covalent enzyme-DNA complex?

tyrosine

coenzymes

organic molecules that provide functional groups for enzyme catalysis

what are coenzymes often from

vitamins

Examples of coenzymes for catalysis

NAD(P)+/ NAD(P)H

Thiamine pyrophosphate TPP

coenzyme A

folate

What are NAD(P)+ and NAD(P)H

oxidation reduction coenzymes

What is the site of reversible reactions on NADPH/NADH

nicotinamide

Do enzymes bind to NADP portion?

no other portion binds enzyme

What kind of coenzyme is Thiamine pyrophosphate?

activation-transfer coenzymes

what is at bottom of trypsin hole

asp

Which part of TPP binds to enzyme vs with decarboxylation of alpha keto acids?

TPP binds to enzyme

thiazolium ring for decarboxylation

What activates the cofactor with TPP

enzyme activates cofactor, acts as base

Catalytic triad for seerine proteiase

Serine, histadine, aspartic acid

Histidine is involved in what is serine protease

acids base

Serine involved with in serine protease

Covalent catalysis, acts as nucleophile to attack peptide bond to make covalent intermediate

Result of cat triad

Amine and carboxylate

what does his act as in serine protease

base and acid

What is generated in step 1 of serine protease

tetrahedral oxyanion intermediate

After step 2 what does histidine act as

acid

What stabilizes the transition state in serine proteases?

hydrogen bonding with backbone amide hydrogens in oxyanion hole to lower EA

Role of metal ions

1. orient substrates for rxn

2. shield and stabilize - charges

   1. donate or accept electrons in redox enzymes

example of metal ions sheild/ stabilize charge

Zn with alcohol dehydrogenase

example of donate and accept electrons in redox with metal ions

Fe in cytochrome P450 enzyme

When can you use enzyme inhibitor as drug

• symptoms caused by excessive metabolite

• abnormal cell growth

• infection from a pathogen

what kind of enzyme inhibitors can durgs be?

reversible or irreversible

How do reversible inhibitors bind to enzyme

non covalent interactions

how do reversible enzyme inhibitors bind

covalent bond between inhibitor and target

What is function of HIV protease

cleaves HIV polyprotein into structural proteins needed for viral maturation

What are charahctersits of HIV protease

aspartic protease

dimeric enzyme

transition state inhibitors

ASP role in catalytic triad

stabilizes his

How does HIV protease vary from serine protease

• serine protease uses Ser-His-Asp to make a covalent enzyme substrate intermediate

• HIV prease uses two Asp residues to activate water to attack peptide bond

how does duranavir inhibit HIV protease?

it is a transition state mimic, so it binds through hydrogen bonds with HIV protease to block viral matration,

function of hydrogen bonds with DRV and HIV protease

stabilize DRV binding to prevent cleaving of polyproteins

5 steps of serine protease mechanism

1. General base catalysis: Oxyanion intermediate forms.

2. General acid catalysis: Covalent intermediate forms.

3. Amine leaving group is released.

4. General base catalysis: Another oxyanion intermediate forms.

5. General acid catalysis: Carboxylic acid leaving group is released.

what is a activation transfer voenzyme

takes place in rxn by binding covalently to a substrate temp

Which AA act as acids?

histidine, asp, glu

which AA act as bases

Lys, Arg, His

Oxidation value of O, C, H

0 takes all

1

2