Ppt 1 Flashcards
Browning in apples is caused by what?
polyphenol oxidase
What is the function of enzymes?
enable biological reactions that proceed at perceptible rates in living organisms
Enzymes in animals, plants, and microorganisms all possess the same what?
Functional classes
In what two way are food enzymes classified?
- Exogenous: added to food for desirable change
2. Endogenous: exist in food affecting the quality (rancidity, browning, etc)
Enzymes can be hydrolyzed by what 3 things?
Acids, bases, low molecular weight peptides (proteases?)
What enzyme breaks down other enzymes? What does it form?
What precipitates out of these reactions?
Proteases - forms amino acids / peptides
Water
What are 3 common protein tests?
buret test, lowry test, BCA test
Enzymes possess what 3 important traits?
- They are proteins
- They are catalysts
- They exhibit selectivity towards substrates
What are coenzymes?
non-protein components that help enzymes carry out catabolic function
Haloenzyme
Complete enzyme with a functional activity; _____.
- For some enzymes, the holoenzymes equal the protein part (ex. trypsin, chymotrypsin)
- For other enzymes, the holoenzyme requires the essential non protein part for functional activity(polyphenol oxidase, peroxidase)
Apoenzyme
- “protein part”
- void of essential cofactor
- inactive enzyme. activation occurs upon binding of an organic or inorganic cofactor
Prosthetic group
- essential non protein part
What is an active site?
Region that binds and transforms substrates into products
What are 3 features of active sites?
- small
- 3D entities
- cleft / crevices
What 2 events happen to the active site during enzymatic reactions?
- binding of S
- transformation
- not all binding leads to transformation but no transformation happens without binding
What is bond specificity? Also called?
enzymes act on compounds with similar bonds.
Also called relative/low specificity
Give 2 examples of bond specificity:
- lipase: acts on ester bonds in lipids/fats
2. Protease: acts on peptide bonds in protein
What is group specificity? Also called?
enzymes act on a group of related compounds. Also called moderate specificity
Give 2 examples of group specificity
- Pepsin: can hydrolyze peptide bonds where there is aromatic amino acids (Phe.,Try., Typ.)
- Trypsin: canhydrolyze where there is basic amino acids (Lys., Arg., Histidine)
What is absolute specificity? Also called?
Enzyme acts on only one single substrate. Also called substrate or high specificity
Give 2 examples of substrate specificity
- Lactase –> Lactose
2. Sucrase –> sucrose
What is stereospecificity? Also called?
Enzyme acts on isomer of a molecule. Also called “high specificity”
Give an example as to why it is necessary to know enzyme specificity for food science?
Starch:
- linear (alpha1–>4)
- branched (alpha1–>6) more sticky
— to make a less sticky product you would need an enzyme to detach the polymers
Enzyme commission numbers - how many?
4 (EC a.b.c.d.)
What does each digit stand for (a.b.c.d.) in Enzyme Commission numbers?
a. group
b. sub-group
c. sub-sub-group
d. numbering
What does the 1st No. ‘a’ denote in EC a.b.c.d.?
‘a’ = type of reaction catalyzed. 6 main types of reactions are distinguished (1-6)
What is E.C. 1?
Oxidoreductase
What do oxedoreductase reactions catalyst? 2 examples
redox rxns
- polyphenol oxidase (PPO)
- glycol oxidase (GOX)
What is E.C. 2?
Transferases
What do transferase reactions catalyst? 1 example
Catalyze the transfer of various groups from one molecule (donor) to another (acceptor)
- Transglutaminase (TEASE)
What is E.C. 3?
hydrolases
What do hydrolases reactions catalyst? 3 examples
Catalyze the cleavage or hydrolysis of larger molecules into smaller molecules with H2O as a co-reactant
- Protease, Lipase, Carboxylase
What is E.C. 4?
Lyases
What do lyase reactions catalyst? 2 examples
Remove groups from substrate; products formed have low molecular weights (usually with unsaturated bonds)
- Pectin
- Lyase
What is E.C. 5?
Isomerases
What do isomerase reactions catalyst? 1 example
catalyze the conversion of molecules into their isomers
- glucose isomerase
What is E.C. 6?
Ligases
What do Ligase reactions catalyst? 1 example
Catalyze the joining together of two or more molecules
- Fatty acyl CoA synthase
Why is enzyme purification necessary?
to remove other undesirable components from the source materials (toxins, other enzymes, biomolecules like DNA)
Enzyme extraction process
raw materials are blended or homogenized in buffer solutions
Enzyme purification
size/charge of enzyme is different. Hydrophobic/hydrophilic effects solubility
What are the 4 types of enzyme purification methods based on size difference?
- dialysis
- ultrafiltration
- centrifugation
- gel filtration
What is special about a dialysis membrane? What is it made up of?
semi-permeable film - normally cellulose with different sizes of pores.
How does dialysis work?
Sample is placed in dialysis bag. Bag is placed in a solvent of more dilute concentration than sample solution. Small molecules in the sample travel through pores to the outside of the bag while the larger ones stay in the bag. When equilibrium is reached, smaller molecules can re-enter the bag. It is possible to change the solvent several times to remove as much of the small molecules as possible.
What is the overall purpose of dialysis?
separate small and large molecules
Is dialysis efficient?
no
Ultrafiltration
uses semi-permeable membrane. Pressure in the form of gas (N2) or vacuum is used to force the small molecules through the pores in the membrane
Centrifugation
Uses centrifugal force to speed up sedimentation of different molecules in a suspension
In centrifugation, what molecules move at a faster rate?
larger/heavier
What happens during gel filtration electrophoresis?
Resin is placed in column, sample applied to the resin. Sample migrates down column, larger molecules pass through spaces & elute faster while smaller molecules enter the pores.
What are the 3 types of enzyme purification methods based on charge difference?
- Ion exchange chromatography
- Electrophoresis
- Isoelectric focusing
What are the 3 types of enzyme purification methods based on solubility differences?
- Isoelectric precipitation
- Salt fractionation
- Solvent precipitation
How can the solubility of an enzyme be characterized at its isoelectric point?
minimal
What happens during isoelectric focusing?
pH of sample solution is adjusted so that the enzyme can precipitate at a pH value corresponding to its isoelectric point.
What are the 3 types of enzyme purification methods based on specific binding sites?
- affinity chromatography
- hydrophobic interaction chromatography (HIC)
- hydrophilic interaction chromatography (HILIC)
Affinity chromatograhphy
- Substances linked in resin have specific binding with particular enzymes
- – substrate, cofactor, inhibitor
- Unbound enzyme will elute from the column at first
- – bound enzyme can be washed out of column by changing the pH or ion strength of the elution buffer
How many purification methods must be used to get an enzyme fully pure?
3 or 4
- electrophoresis & isoelectric focusing
- activity testing
- chromatographic behaviour
