Aim

The aim of our modelling is to be able to estimate approximate yields of vanillin when x amount of tyrosine is entered into the pathway whilst also showing approximate times.

This also allows the possibilty to choose other strains of the enzymes that we have also used within our different constructs.

By being able to choose between other strains we are more able to optimize the overall vanillin yield, whilst also being able to see any potential bottlenecks within the substrates by viewing increased concentrations throughout the reaction.

How

The model employs Michaelis-Menten kinetics to simulate the enzymatic reactions occurring at each step of the pathway. The concentration of substrates and products is calculated over time, this allows the ability to see the total vanillin at the end of the reaction aswell as the concentrations at every second of the reaction.

$Super E. coli$

The model also uses accurate conversion rates for each strain, allowing the ability to see the most efficient strains aswell as any bottlenecks they may cause. The enzyme kinetics and conversion rates have been obtained from similar experiments found in literature that have used the same strain.

Limitations

Optimal Temperature

The model assumes that each strain is constantly at its 'optimal temperature range' as different strains that we have used have have different optimal temperature ranges for example Pseudomonas putida KT2440 is optimal at 37°C where as Streptomyces sp. V-1 [7].

This limitation will slighly impact the theoretical model results to the actual results, as not all strains will be able to be at optimal temperature range.

Byproducts
The model is unable to show the byproducts that are also produced during the reactions, this would be useful in cases for example, when large amounts of L-Dopa is produced which creates a dark pigmentation as we saw in our results.

Tyrosine to Vanillin Calculator

TAL:

(Tyrosine to p-Coumaric Acid)

C3H:

(p-Coumaric Acid to Caffeic Acid)

COMT:

(Caffeic Acid to Ferulic Acid)

FCS:

(Ferulic Acid to Feruloyl-CoA)

ECH:

(Feruloyl-CoA to Vanillin)

Initial Tyrosine Amount (µM): µM

Enzyme Kinetics

TAL

Substrate	Product
Tyrosine	p-Coumaric Acid

(SAM8)

Parameter	Value
k_cat	0.015 s^-1 [1]
K_m	15.5 µM [1]
Yield Efficiency	87.33% [2]

C3H

Substrate	Product
p-Coumaric Acid	Caffeic Acid

(SAM5)

Parameter	Value
k_cat	4.2385 s^-1 [3]
K_m	432.0 µM [3]
Yield Efficiency	75% [3]

COMT

Substrate	Product
Caffeic Acid	Ferulic Acid

(ARA)

Parameter	Value
k_cat	0.03319 s^-1 [9]
K_m	43.0 µM [9]
Yield Efficiency	70.33% [10]

FCS

Substrate	Product
Ferulic Acid	Feruloyl-CoA

(Pseud)

Parameter	Value
k_cat	0.074 s^-1 [4]
K_m	43.0 µM [4]
Yield Efficiency	86% [8]

(STR)

Parameter	Value
k_cat	67.7 s^-1 [6]
K_m	350.0 µM [6]
Yield Efficiency	97.1% [6]

ECH

Substrate	Product
Feruloyl-CoA	Vanillin

(Pseud)

Parameter	Value
k_cat	0.4 s^-1 [5]
K_m	75 µM [5]
Yield Efficiency	86% [8]

(STR)

Parameter	Value
k_cat	77.36 s^-1 [5]
K_m	0.4 µM [5]
Yield Efficiency	97.1% [6]

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