ENZYME BIOCHEMISTRY MICHAELIS- MENTEN THEORY OPT 4 MLT 435 NAME NUR FATIHAH BINTI SAIFUL EFFENDY NAYLI NUR FARZANA BINTI MOHD SHAHARUZZAIN NUR ALIYA DANISHA BINTI MOHD JAMIL NUR IFFAH BINTI MOHAMMAD ZAIBI HUSNA NAJIHA BINTI HUSSAINY ID NUMBER 2022830304 2022611716 2022868186 2022622422 2022879576
E+S ES E+P v = Vmax [S] Km + [S] Vmax = maximum velocity at maximum substrate concentration Km = Michaelis constant S = concentration of substrate S Km Vmax 2 Vmax Reaction velocity, V Substrate concentration [S] Relationship between kinetic parameters Vmax and Km can be seen in this graph of the Michaelis-Menten equation's estimated reaction velocity as a function of substrate concentration. At a Berlin hospital, Maud Leonora Menten, a Canadian biochemist and organic chemist studied enzyme kinetics with Leonor Michaelis, a biochemist of German descent. Michaelis and Menten were able to express mathematically the relationship where each enzyme not only has a specific substrate, but when there are adequate amounts of that substrate present, it also changes chemically at a rate specific to that enzyme. WHO INVENTED MICHAELIS-MENTEN EQUATION? Introduction MICHAELIS-MENTEN EQUATION - REACTION BETWEEN VELOCITY TO SUBSTRATE CONCENTRATION In a system where a substrate S binds reversibly to an enzyme E to form an enzyme-substrate complex ES, which subsequently reacts irreversibly to produce a product P and regenerate the free enzyme E. MICHAELIS-MENTEN EQUATION
An assumption made in the derivation of the Michaelis-Menten kinetics which states that the binding of substrate to the enzyme produce enzyme-substrate complex occur very quickly compare to the complex catalyse product and that these processes are in equilibrium. This means that the rate of formation of the enzyme-substrate complex is equal to the rate of its breakdown, and the concentration of the enzymesubstrate complex remains constant over time. This assumption allows for the simplification of the kinetics equations by assuming that the rate of the enzymatic reaction is determined by the concentration of the enzyme-substrate complex and not by the individual concentrations of the enzyme and substrate. This assumption is valid for many enzymes, but it may not hold true for enzymes that bind very slowly to substrate or for enzymes that have multiple steps in their catalytic mechanisms. Assumptions 1. RAPID EQUILIBRIUM ASSUMPTION - HENRI-MICHEALIS-MENTEN EQUATION The substrate binding process can be derived by equilibrium dissociation constant (Kd): Michealis constant (Km) is equal to Dissociation constant (Kd) in this assumption. Kd = Km
The Michaelis-Menten kinetics assumes that the system is at steady state, meaning that the rate of formation of the enzyme-substrate complex is equal to the rate of its breakdown. Steady-state assumption is the first important assumption involve in Briggs and Haldane's derivation. In other words, the concentration of the enzyme-substrate complex remains constant over time, and the rate of the enzymatic reaction is determined by the concentration of the complex. Assumptions 2. STEADY-STATE ASSUMPTION k : rate constant - k1<< k-1 >> kcat Assuming that [P] = 0 High concentration of substrate, [S] added to low concentration of enzyme, [E ] forming complex ES. ES is at steady state and will be constant to catalyse E+P. another method to develop and expression of free enzyme [E} by assuming the concentration of [ES] is constant during binding reaction : This assumption allows for the simplification of the kinetics equations, the determination of the maximum rate of the reaction (Vmax), and the substrate concentration at which the reaction is half-maximal (Km) which are important parameters to characterise the enzyme's kinetic properties.
Derivation of Michaelis Menten Equation THIS EQUATION BASICALLY DESCRIBES WHAT HAS TAKEN PLACE INSIDE A BEAKER ONCE WE ADD THE SUBSTRATE. As we form the ES complex, the rate constant is k1. When it begins to dissociate back into E & S the rate constant will be k-1. As the complex (ES) formed, the enzyme will begin to transform that substrate (S) into product (P) making the rate constant as k2 and thus producing enzyme (E)and product (P) in its individual form. As it go in reverse, the rate constant is k-2. To simplify the equation, we can study the reaction at the beginning, that is when t ≈ 0 and V=Vo. At this moment in time, very little product will form and so the reverse of the product (P) formation reaction k-2 will be negligible. Hence: The enzyme-substrate complex intermediate is our starting point. The rate (Vo )at which the substrate is transformed to a product (as picture above) is equal to the product of rate constant k2 multiplied by the intermediate enzymesubstrate complex. Thus:
2 However, the reaction ES could also dissociate and reform E and S which is equal to the product of the reverse reaction k-1 multiplied by ES. Plus, our ES complex not only dissociate going this way, but there’s also a probability that it will dissociate going through k2 path. So, we will have k2 multiplied by ES. As a result, these two ways will give us summations of ES complex breakdown equation: 3 To form ES, rate constant k1 is multiplied by the concentrations of each E and S Derivation of Michaelis Menten Equation 1 4 To get onto the next step, we need to assume the steady-state condition which the concentration of the enzyme-substrate complex will remain constant. So, the rate of formation is equal to the rate of dissociation. Instead of using the ratio of k-1 + k2 divided by k1, we will set it to a new ratio which is Km, also known as Michaelis constant. Hence: 5
6 Now, we will substitute 6 & 7 into 5 : 8 7 Substitute 8 into 1 : 9 Finally substitute into to produce the final Michaelis-Menten equation 9 At the beginning of reaction, when t=0, the substrate total is equal to the [substrate that is not bound to the active site] plus [substrate that is bound to the active site]. However, when we add a substrate, the ES quantity is so much smaller than S quantity hence it is negligible: Essentially, what we assume just now can’t be applied for the enzyme case. Thus, the equation is leave as it is. DERIVATION OF MICHAELIS MENTEN EQUATION Recall that the reaction reaches a maximal velocity, Vmax when all of the enzymes’ active sites are occupied. 10
Graph& Explanation Michaelis-Menten graph plotting consists of : GRAPH EXPLANATION Y-axis = Rate of reaction / Reaction velocity (V) X-axis = Substrate concentration [S] Vmax = Maximum reaction rate Km = Michaelis constant First order kinetics Zero order kinetics The Michaelis-Menten graph explains about the relationship between the reaction velocity and the substrate concentration. There are 2 parts in the graph that can be interpreted : → The reaction velocity is directly proportional to the substrate concentration indicating that the enzyme binds with the substrate continuously. → In this phase, there will be the Km, Michaelis constant where the substrate concentration is at half of the reaction velocity (Vmax/2). → Km is used to measure the affinity of the enzyme for its substrate. → The lower the value of Km, the more efficient the enzyme to carry out its function in a lower substrate concentration. → Plateau has reached, the increasing of the substrate concentration does not affect the reaction velocity as it has reached the maximum rate of reaction, Vmax. → Vmax is the maximum reaction velocity where all the enzyme active sites is already saturated with the substrate. → There is no free enzyme left to bind with the remaining increasing substrate.
A A B B A B A B A B A B A B A B A B A B A B A B Significance of Michaelis Menten CAN PREDICT WHETHER THE CELL NEEDS MORE ENZYMES OR MORE SUBSTRATE TO SPEED UP THE ENZYMATIC REACTION. IF AN ENZYME CAN CATALYSE A REACTION WITH TWO SIMILAR SUBSTRATES (E.G., GLUCOSE AND FRUCTOSE) IN THE CELL, IT WILL PREFER THAT SUBSTRATE FOR WHICH THE ENZYME HAS LOWER KM VALUE GIVES AN APPROXIMATE MEASURE OF THE CONCENTRATION OF SUBSTRATE OF THE ENZYME IN THAT PART OF THE CELL WHERE REACTION IS OCCURRING.
REFERENCES MLT 435 University of Washington. (2019b). Michaelis-Menten Kinetics and Briggs-Haldane Kinetics. Washington.edu. https://depts.washington.edu/wmatkins/kinetics/michaelis-menten.html Leonor Michaelis and Maud Leonora Menten. (2016, June 1). Science History Institute. https://www.sciencehistory.org/historical-profile/leonor-michaelis-and-maud-leonoramenten Maud Leonora Menten | Canadian biochemist and organic chemist. (n.d.). Encyclopedia Britannica. https://www.britannica.com/biography/Maud-Leonora-Menten Le, H., Algaze, S., & Tan, E. (2019, June 5). Michaelis-Menten Kinetics. Chemistry LibreTexts. https://chem.libretexts.org/Bookshelves/Biological_Chemistry/Supplemental_Modules_(Biolo gical_Chemistry)/Enzymes/Enzymatic_Kinetics/Michaelis-Menten_Kinetics Surti, F. (2022, November 13). Enzyme kinetics - structure - function - michaelis-menten kinetics. TeachMePhysiology. Retrieved from https://teachmephysiology.com/biochemistry/moleculesand-signalling/enzyme-kinetics/ Team, P. W. (2014, September 1). Michaelis-menten equation - interactive graph. PhysiologyWeb. Retrieved from https://www.physiologyweb.com/calculators/michaelis_menten_equation_interactive_graph.h tml Km vs Kd - the difference between Michaelis and dissociation constants. The Science Snail. (n.d.). Retrieved January 27, 2023, from https://www.sciencesnail.com/science/archives/02- 2019 Michaelis-menten (steady-state) kinetics the Michaelis-Menten Model K ... (n.d.). Retrieved January 26, 2023, from https://www.chem.ucla.edu/~rebecca/153A/MMkinetics.pdf Ziemke, T. (2022, December 20). Steady State approximation. ChemTalk. Retrieved January 27, 2023, from https://chemistrytalk.org/steady-state-approximation/ 1. 2. 3. 4. 5. 6. 7. 8. 9.