Enzymes Flashcards
enzyme specificity
Absolute specificity
Protease -> protein
Lipase-> lipida
Group specificity
Enzyme act on a group of compounds
Stereospecificity
Enzyme that cat on optimal isomer
Race made and epimerase
Racemase- act on molecule with 1 chiral carbon
Epimerase - multiple chiral carbon
Enzyme purification is based on what
Size difference
Solubility diff
Charge diff
Selective absorption
Size diff is based on what
Dialysis
Ultrafiltration
Centrifugation
Gel filtration
Dialysis what is it and what is the problem
Enzyme is placed in a dyalisys bag and small molecule can diffuse bc of the small pores
Problems: every time there is an equilibrium dialysis cannot completely move the molecule
Ultrafiltration
Apply pressure in one side and force smaller molecule through membrane
Centrifugation
Larger molecules settle down faster
Gel filtration
Larger ones go down more quickly
Separation based on solubility is based on
Isoeletric preciptation
Salt fractionation
Solvent precepitation
Isoeletric preciptation
PH is adjust u t’il the net charge is neutral. PH ranges, diff enzymes have diff pIs and this is used to separate enzymes
Salt fractionation
Salts can alter charges on enzymes molecules
If you add salt more soluble
Solvent preciptation
Acetone or ethanol are used or decrease water activity you need to chill solvent to 0-4 degrés before adding the enzyme to prevent enzyme denaturation
Separation based on charge differences are based on
Ion exchange chromatography
Electrophoresis
Isolectric focusing
Ion exchange chromatography
Cation exchanger when + binds to -
Anion is the inverse
Electrophoresis
Based on size and charge
The larger the molecule the faster
Isoélectrique focusing
We genetate pH gradient in the gel by using ampholytes
Separation based on specific binding based on
affinity chromatography and hydrophobic interaction chromatography
Affinity chromatography
Ligand are bound to enzymes with resin
The principle of affinity chromatography is that the stationary phase consists of a support medium (e.g. cellulose beads) on which the substrate (or sometimes a coenzyme) has been bound covalently, in such a way that the reactive groups that are essential for enzyme binding are exposed. As the mixture of proteins is passed through the chromatography column, those proteins that have a binding site for the immobilised substrate will bind to the stationary phase, while all otter proteins will be eluted in the void volume of the column.
HIC: hydrophobic interaction chromatography
Bind hydrophobic material to enzymes after the unbound material are eluted and the bound material are next removed by pH change
Enzyme purity is tested by
Test for homeogeonity
Chromatographic behaviour
Active testing
Isolectric fusing
Test for homogeonity
Make sure is homogenous by electrophoresis. Serial band = pure
Chromatography behaviours
Take ans and get various peaks if they are symmetrical is oure
Activity testing
Use diff substrates to know which enzyme is present. If it is more than one so it is not pure
Put the enzyme groups in order
1) oxidoreductase
2) transferase
3) hydrolyse
4) lyase
5) isomerase
6) ligase
7) translocase
True or false
Enzyme facilitate reactions by reducing energy barrier
True
What is the effect of temperature on catalysis
The kinetic energy will increase until the optimum temperature after it will be degraded
Effect of pH on catalysis
At externes pHs the enzyme will be denatured and there is an Ph optimum around ph 7
What are the 3 types of enzyme inhibition
Competitive
Non competitive
Uncompetitive
Competitive inhibition
The inhibitor will bind to the enzyme and take the place of the substrate. We can increase substrate concentration to overcome this problem
Non competitive inhibition
The inhibitor will bind to another place on the enzyme and it cannot make products. It can overcome by doing dialysis or gel filtration to remove the inhibitors
Uncompetitive inhibiton
The substrate will bind to the active site and it will open up another place so the inhibitor can bind.
What is the beneficial aspects of food processing
Natural, non toxic
Specificity
They are effective under mild reactions
Not expensive
Easy to control
What are some deleterious effects of food processing
Lipase/ phospholipase can cause hydrolysis of these compounds into free fatty acids that can form various oxidized compounds that can make off flavours
In canned foods if the caning process is not complete, the heat stable enzymes ( protease) may survive the canning and continue with protéolytique even during storage in the can
Cheeses proteins lysis can make bitter peptides and alter the taste
What enzymes does cheese use and why
Enzymes will do the same function. The enzymes that can be presents are rennin, rennet, pepsin or chymosin
In cheese there is casiers bc of the mild. Enzyme can cleave the Phe(105)-met(106) peptide bond in the k-casein to enable curdling to happen. But when you add the enzyme they form a low molecular weight peptides that will remain in the solution that is whey
What enzymes are found in winemaking and brewing and why
Amylase : break down carb to form glucose, the glucose is fermented by yeast and produce CO2 and alcohol
Protease : they are added to break down proteins in beverage into low molecular weight peptides and amino acids that remain in solution at refrigerated temperatures it will prevent precipitation of proteins
What enzymes are used in baking and why
Amylase : breaks down starch into glucose that are fermented by yeast to make CO2 that enables the volume to swell
Lipase: produce volatile compounds
Lipoxidase : bleach the color of flour from yellow to white
What are the enzymes used in meat industry and why
Protease : break down the large myofibrillar proteins to form low molecular weight peptides and free amino acids and it results in a more tender and modified taste
Lipase : flavor
Enzymes can also be added to add smaller molecules into a buffer one in the fish meat industry. Which enzyme is it and why do we put it
Transglutamine ( TGAse) : they form isopeptide bond between NH2 groups in the side chain of lysine and glutamine.
The isopeptide bond are covalent bond because the products can be larger and though
What are the chemical reactions in foods catalyses by
Proteases, carbohydrates and lipolytic enzymes ( hydrolases)
Chemical reaction if oxidoreductase
I’m chemical reactions of protease there is 2 groups what is it and what they do
( endoprotease va exo)
1) endoprotease
Cleave provide bond randomly in the protein molecule
It impacts texture and solubilisation
2) exoprotease
Cleave peptide bond at the terminal protein and impact flavour
What are the 2 groups of carbohydrase
Starch and cellular
Starch has an alpha 1-4 and alpha 1-6 linkages and cellulose has beta 1-4 linkage ( linear structure) and so cellulose can be more compacted than starch bc it is branched but it is more resistance to hydrolysis
Is cellulose more resistant than starch
Yes because it can be more compacted because if it’s linkage ( alpha 1-4) because starche has a branched
What is the most important function of lipase
Lipolysis bc it brings hydrology’s to the major fat ( triaglylcerides) to form glycerol and free fatty acids
What lipolysis do
Texture and flavour changes
Texture bc of of the larger molecule becoming the free fatty acids and glycerol
Flavor bc of the oxidation of free amino acids
Is lipase desirable or not
Depends on the food. It is not desirable in meats and fish but desirable in cheeses and chocolates
What is the role of amino acid oxidase
Form Keto acids, ammonia and H2O2. It is made when a person doesn’t have a lot of carbs in the diet
Role of PPOs ( polyphenol oxidase)
They catalyze the oxidation of phenolic compounds to form dark coloured compounds ( melanidins)
Is melanoidin ( dark compounds ) desirable
Yes in chocolate tea coffe but not in Crustacea, vegetable
Tile of glucose oxidase
Oxidation of glucose to form gluconic acid and H2O2
Role of catalase and peroxydase
Breading of H2O2 to form less reactive H2O and O2
Catalase is specific for H2O2 and peroxidase for both
Why catalase and peroxidase are important
Prevent oxidative damage
What is the role of ça then oxidase
Catalase the hydroxylation of hypoxanthine to xanthine and xanthine is further oxidize and it will produce uric acid which is not good because it decreases the quality effects
What are some examples of enzymatic modification of foodstuff
Change in solubility, functional properties and flavor changes
What are some industrial application of proteases
1) removal of bitterness
2) modification of milk and whey proteins
3) hydrolysis of what gluten for use as flavoranys
4) modification of collagen and gelatin ( modified collagen )
5) alcoholic beverage
6) baking
7) recovery of scrap protein form offal bones and blood
What are other less known application of enzymes
Removal of dental plaque with toothpaste containing des transes
Elimination of hair with keratinase
Solubilisation of cold tea solid with tanase
Synthesis of protein like molecule
Methods for controlling enzymatic reactions in food
Traditional :
Based on temperature
Based on water activity
Based on chemicals
What are the Nobel methods for controlling enzymatic reactions
High pressure treatments
Enzyme treatments ( killer enzyme)
Enzyme inhibitors ( inhibit protéolytique in fish flesh or meat prevent mushiness)
Chemical modification : side chains of enzymes molecules may be modified by attaching groups to modify the reactivities of the enzymes
Ionizing radiation : cause radiolysis of H2O molecules and the product of radiolysis of water can bind to the enzyme molecule and cause inhibition
