Add to your concept map Write all the key words from last lesson – leave some space so you can add more in coming lessons (2 word trigger meaning with it) Join different word – write how they are joint on the line .... Induced fit Activation energy Orientation Affinity Metabolism – chemical reactions 2 types Catabolism –breakdown , energy made

Regulation of metabolic pathways by the action of enzymes

How does your body prevent “burnout”? Clue 

Gene Expression of Enzymes Some metabolic pathways are only required to operate under certain circumstance Prevent wasted resources by “switching on/off” of genes that code for the enzymes controlling each of the stages

Learning Outcomes Control by gene expression - give an example of lactose metabolism being switched by presence/absence of lactose (Lac-operon proposed by Jacob-Monod) Control by signalling molecules (adranalin) Control by inhibitors or competitors

Genes & Escherichia Coli (E.Coli) E.Coli preferred food source is glucose Lactose is disaccharide made up of glucose and galactose. E.Coli has gene for β-glactosidase which digests lactose to glucose and galactose Gene for β-glactosidase in E.Coli on switched on when lactose present – off usually by the repressor upstream of lac operon Process of switch on is called enzyme induction -

Operon consists of; Regulator consists of; 1 or more (3 here) structural genes which code for enzyme operator region controls switching on/off of Promoter site for RNA polymerase to bind to Regulator consists of; Promoter site for RNA polymerase to bind to Gene for making repressor protein phET Animation

When lactose present ...... Up to 4:40mins

Gene action hypothesis was put forward by Jacob-Monod Confirmed by many experimental experiments – one of which you will complete next lesson ..... Jacob, Monod & Lwoff © NobelPrize.org

Regulation by signal molecules Intracellular Signals ;Lactose is an example of a signal molecule from the cells environment Extracellular Signals; Epinephrine (Adrenalin)is also a signal molecule – this time outside the liver cells (in blood stream by adrenal glands) – but it triggers conversion of glucose from stored glycogen in liver cells.

Regulation by inhibitors 2 types; Competitive inhibitors Non-competitive inhibitors Compete for the active site Directly compete Overcome by increasing substrate concentration Attached to non-active site (allosteric), but this changes the shape of the enzyme molecule. Indirect change Not overcome

As shown below, the competitive inhibitor can be challenged by increasing the substrate concentration – as more enzyme sites will contain substrate not inhibitor. No effect on non-competitive inhibitor when substrate concentration is increased.

1 Enzyme many active sites ... Some enzymes have several polypeptide subunits with their own active sites. Can be either active or inactive Can change shape if regulator molecule (reg) bound to allosteric (non- active site) If reg activato rthe enzyme active and enzyme activity stimulated, rate of reaction If reg non-competitive inhibitor then inactive state and no enzyme activity rate of reaction

Regulation by products Final method of regulation is by end-product feedback inhibition Guess what you think it is – tell your partner Come up with a collaborative idea

End-product Feedback Inhibition The end product acts as an inhibitor, feedback to the pathway. This is then binds to an earlier enzyme that catalyses the reaction earlier in pathway The end product then inhibits this enzyme Causing the slowing down of conversions Called negative feedback control; finely tuned in order not to accumulate intermediates/final products and prevent wasteful conversions! The control of metabolic pathways by feedback inhibition where an end product binds to an enzyme that catalyses a reaction early in the pathway.

Demonstrate your knowledge Use the cards and place them into different categories – you choose categories! Metabolic pathways are controlled by the presence or absence of particular enzymes. Enzymes always act individually. Competitive inhibition can be reversed by increasing the substrate concentration.

Review Complete this extended response in 20 minutes; Describe the 'induced fit' model for enzyme activity and explain how enzyme activity may be regulated.

Swap with someone on another table and peer assess Use the markscheme .....

b-galactosidase

b-galactosidase

Competitive inhibition

Non-competitive inhibition

Non-competitive inhibition

Effect of [substrate] Competitive inhibitor Non-competitive inhibitor Increasing [substrate] displaces inhibitor from active site Non-competitive inhibitor Increasing [substrate] has little or no effect on enzyme activity

Method Carry out reaction Without inhibitor at low [S] (ONPG) In presence of inhibitor Galactose Iodine [I] chosen to completely inhibit reaction and look at increasing [S]

Colorimeter Direction of Beam R = Reference T = Test

1 2 - 1.0 0.25 0.75 3 0.5 4 5 Cuvette No 20% galactose in buffer (cm3) ONPG stock solution (cm3) Buffer (cm3) ONPG x 20 dilution Absorbance 1 2 - 1.0 0.25 0.75 3 0.5 4 5

I = Galactose (steps 6-7) Prepare test tubes Each tube contains 2 cm3 galactose (I) Tubes 1 – 5 contain increasing [S] NOTE – diluted (x 20) ONPG into tube 1, ONPG stock and buffer into tubes 2 – 5 Mix, place in cuvette  colorimeter, reference colorimeter Return to test tube Add 0.5 cm3 diluted enzyme, start timer & mix, add to cuvette, read Absorbance after 2 min

Step 10 Add buffer and diluted (x 20) ONPG to test tube, mix and reference colorimeter Add 0.5 cm3 diluted enzyme, start timer, mix read in colorimeter after 2 min Compare with results after step 5.

Step 10 Mix diluted buffer and diluted ONPG in test tube, zero colorimeter Add diluted enzyme Read absorbance after two minutes Compare with results after step 5.

Results

Results