Michaelis-Menten Kinetics

Slides:



Advertisements
Similar presentations
Enzyme Kinetics C483 Spring 2013.
Advertisements

Biochemistry 2/e - Garrett & Grisham Copyright © 1999 by Harcourt Brace & Company Chapter 14 Enzyme Kinetics to accompany Biochemistry, 2/e by Reginald.
Biochemistry 2/e - Garrett & Grisham Copyright © 1999 by Harcourt Brace & Company Enzyme Kinetics.
Kinetics: Reaction Order Reaction Order: the number of reactant molecules that need to come together to generate a product. A unimolecular S  P reaction.
Enzyme Kinetics, Inhibition, and Control
Enzyme Kinetic Zhi Hui.
Chapter 7 Chem 341 Suroviec Fall I. Introduction The structure and mechanism can reveal quite a bit about an enzyme’s function.
Enzyme Kinetics. Rate constant (k) measures how rapidly a rxn occurs AB + C k1k1 k -1 Rate (v, velocity) = (rate constant) (concentration of reactants)
Medical Biochemistry, Lecture 24
Nonlinear pharmacokinetics
Enzyme Kinetics and Catalysis II 3/24/2003. Kinetics of Enzymes Enzymes follow zero order kinetics when substrate concentrations are high. Zero order.
Enzyme Kinetics: Study the rate of enzyme catalyzed reactions. - Models for enzyme kinetics - Michaelis-Menten kinetics - Inhibition kinetics - Effect.
Chapter 12 Enzyme Kinetics, Inhibition, and Control Chapter 12 Enzyme Kinetics, Inhibition, and Control Revised 4/08/2014 Biochemistry I Dr. Loren Williams.
Inhibited Enzyme Kinetics Inhibitors may bind to enzyme and reduce their activity. Enzyme inhibition may be reversible or irreversible. For reversible.
ENZYME KINETIC M. Saifur R, PhD. Course content  Enzymatic reaction  Rate of Enzyme-Catalyzed Reactions  Quatification of Substrate Concentration and.
Nonlinear Pharmacokinetics
Enzyme kinetics Why study the rate of enzyme catalyzed reactions? Study of reaction rates is an important tool to investigate the chemical mechanism of.
CH13. Enzymes cXXkcZ2jWM&feature=related.
Chapter 6.3: Enzyme Kinetics CHEM 7784 Biochemistry Professor Bensley.
Chapter 5 (part 2) Enzyme Kinetics.
Lecture 6: Kumar Measuring enzyme activity 1. Effect of pH on enzyme activity 2.
Why study enzyme kinetics?  To quantitate enzyme characteristics  define substrate and inhibitor affinities  define maximum catalytic rates  Describe.
Rules for deriving rate laws for simple systems 1.Write reactions involved in forming P from S 2. Write the conservation equation expressing the distribution.
Chapter 5 (part 2) Enzyme Kinetics. Rate constant (k) measures how rapidly a rxn occurs AB + C k1k1 k -1 Rate (v, velocity) = (rate constant) (concentration.
Picture of an enzymatic reaction. Velocity =  P/  t or -  S/  t Product Time.
Paul D. Adams University of Arkansas Mary K. Campbell Shawn O. Farrell Chapter Six The Behavior of Proteins:
The Michaelis-Menton Model For non-allosteric enzymes, the most widely used kinetic model is based upon work done by Leonor Michaelis and Maud Menton For.
NON LINEAR PHARMACOKINETICS Dr. Muslim Suardi, MSi., Apt. Faculty of Pharmacy University of Andalas 2013.
Michaelis-Menten kinetics
Nonlinear Pharmacokinetics
Process Kinetics Lecture 1 Mahesh Bule 4/27/2017
Enzyme Kinetics I 10/15/2009. Enzyme Kinetics Rates of Enzyme Reactions Thermodynamics says I know the difference between state 1 and state 2 and  G.
Rmax and Km (26.4) Constants from Michaelis-Menten equation give insight into qualitative and quantitative aspects of enzyme kinetics Indicate if enzyme.
R max and K m (26.4) Constants from Michaelis-Menten equation give insight into qualitative and quantitative aspects of enzyme kinetics Constants – Indicate.
6.1 A Brief Look at Enzyme Energetics and Enzyme Chemistry Converting substrates to product requires intermediate states – Intermediates are less stable.
ENZYMES 2.
Enzyme Kinetics Sadia Sayed. What is Enzyme Kinetics?  Kinetics is the study of the rates at which chemical reactions occur  Then what is Enzyme Kinetics?
Lecture 5:Enzymes Ahmad Razali Ishak
Interpretation of Michaelis Menten Equation. Michaelis-Menten  Graphically representation:
Enzyme kinetics & Michaelis-Menten Equation Abdul Rehman Abbasi MSc Chemistry Semester – I Preston University Isb.
Enzyme Kinetics Enzyme Kinetics:
Basic enzyme kinetics Concepts building:
Enzyme Kinetics Bwahahahaha!
Basic enzyme kinetics Concepts building:
Enzymes.
Determination of the Kinetic activity of beta-fructofuranosidase and the Mechanism of Inhibition by Copper (II) Sulfate.
Enzyme Kinetics provides Insight into
Bioreactors Engineering
Lecture 15 Chemical Reaction Engineering (CRE) is the field that studies the rates and mechanisms of chemical reactions and the design of the reactors.
Enzymes II Dr. Kevin Ahern.
Enzymes Packet #13 Chapter #8 Friday, November 23, 2018.
ENZYME INHIBITION.
Enzymes II:kinetics Dr. Nabil Bashir.
Thermodynamics Determines if the reaction is spontaneous (does it occur). Does not tell us how fast a reaction will proceed. Catalysts (enzymes) can lower.
Lecture 15 Chemical Reaction Engineering (CRE) is the field that studies the rates and mechanisms of chemical reactions and the design of the reactors.
Chapter Three: Part Two
Chapter 6 CHM 341 Fall 2016 Suroviec.
The Vmax and Km values of a certain enzyme can be measured by the double reciprocal plot (i.e., the Lineweaver-Burk plot).
13 part 2 Enzyme kinetics 酵素動力學 溫鳳君0993b303 姜喆云0993b039.
(BIOC 231) Enzyme Kinetics
Chapter Three: Part Two
Lecture 8 Enzyme Kinetics
Enzyme Kinetics Nilansu Das Dept. of Molecular Biology
Lecture 15 Chemical Reaction Engineering (CRE) is the field that studies the rates and mechanisms of chemical reactions and the design of the reactors.
Lecture 9 Web: pollev.com/ucibio Text: To: 37607
Enzyme Kinetics Velocity (V) = k [S]
Enzymes Function and Kinetics.
Lecture 15 Chemical Reaction Engineering (CRE) is the field that studies the rates and mechanisms of chemical reactions and the design of the reactors.
23.4 Chain polymerization Occurs by addition of monomers to a growing polymer, often by a radical chain process. Rapid growth of an individual polymer.
Presentation transcript:

Michaelis-Menten Kinetics Presented by Avinash Kodoori M.Pharm I semester Dept. of Pharmaceutics, Univ. College of Pharmaceutical Sciences, Kakatiya University, Warangal.

Contents Linear and Non linear kinetics Causes The Michelis Menten kinetics Introduction Assumptions Derivation Important conclusions Understanding Km and Vmax Determination of Km and Vmax In vitro , Lineweaver-Burk plot In vivo Applications

Effect of dose on riboflavin urinary recovery when given on an empty stomach

Effect of dose on ascorbic acid absorption. April 7, 2019

Steady-state Vitamin C plasma concentration as a function of dose in 13 female subjects receiving doses from 30 to 2,500 mg.

CAUSES OF DOSE DEPENDENT KINETICS Drug GI absorption Saturable transport in gut wall Riboflavin, L-dopa Intestinal metabolism Propanolol Distribution Saturable plasma protein binding Salicylic acid , phenytoin Cellular uptake Methicillin Renal elimination Active secretion p-amino hippuric acid Tubular reabsorption Riboflavin, ascorbic acid Metabolism Saturable metabolism Phenytoin, valproic acid Metabolite inhibition Diazepam

The Michaelis-Menten Kinetics Louis Michaelis and Maude Menten's theory (1913) Applies to – Enzymes - Carriers - proteins Related terms - Dose dependent kinetics - Saturation Kinetics - Capacity limited kinetics

Saturation Kinetics/ Capacity limited kinetics

Assumptions It assumes the formation of an enzyme- substrate complex It assumes that the ES complex is in rapid equilibrium with free enzyme Breakdown of ES to form products is assumed to be slower than 1) formation of ES and 2) breakdown of ES to re-form E and S

Derivation k-2 For the reaction E + S ES P + E the reverse reaction can be neglected at the beginning (initial rate with [P]=0) k-2

1) The rate of formation of the product v = k2[ES], 2)[Enzyme]total = [E]t = [E] + [ES] 3) At steady state: d[ES]/dt = 0 4) KM = (k-1 + k2)/k1

The rate of formation of ES is given as Assume steady state: d[ES]/dt = 0 So: k1[E][S] = k-1[ES] + k2[ES]

[ES] = ([E][S])/KM Rearranging we get [ES] = (k1/(k-1 + k2))[E][S] Substituting (KM = (k-1 + k2)/k1): [ES] = ([E][S])/KM KM[ES] = [E][S] Substituting ([E] = [E]t - [ES]): KM[ES] = [E]t[S] - [ES][S]

Rearranging: [ES](KM + [S]) = [E]t[S] So: The rate of formation of the product is given as

the maximum rate is given as hence

The Michaelis Mentens Equation

At [S] « Km, Vo is proportional to [S] At [S] » Km, Vo = Vmax

Important Conclusions of Michaels - Menten Kinetics when [S]= KM, the equation reduces to when [S] >> KM, the equation reduces to when [S] << KM, the equation reduces to

The "kinetic activator constant" Understanding Km The "kinetic activator constant" Km is a constant Km is a constant derived from rate constants Km is, under true Michaelis-Menten conditions, an estimate of the dissociation constant of E from S Small Km means tight binding; high Km means weak binding

The theoretical maximal velocity Understanding Vmax The theoretical maximal velocity Vmax is a constant Vmax is the theoretical maximal rate of the reaction - but it is NEVER achieved in reality To reach Vmax would require that ALL enzyme molecules are tightly bound with substrate Vmax is asymptotically approached as substrate is increased

The dual nature of the Michaelis-Menten equation Combination of 0-order and 1st-order kinetics When S is low, the equation for rate is 1st order in S When S is high, the equation for rate is 0-order in S The Michaelis-Menten equation describes a rectangular hyperbolic dependence of v on S

a)Invitro determination Lineweaver – Burk Double Reciprocal Plots It is difficult to determine Vmax experimentally The equation for a hyperbola can be transformed into the equation for a straight line by taking the reciprocal of each side

The formula for a straight line is y = mx + b A plot of 1/V versus 1/[S] will give a straight line with slope of KM/Vmax and y intercept of 1/Vmax Such a plot is known as a Lineweaver-Burk double reciprocal plot

Lineweaver-Burk plot (double-reciprocal)

Hanes–Woolf plot invert and multiply by [S]: Rearrange

b. In Vivo Determination Vmax Km K0 K0/Css

JB is an 18 year male receiving phenytoin for prophylaxis of post-traumatic head injury seizures. The following steady state concentrations were obtained at the indicated doses: Dose (mg/d) Css (mg/L) 100 3.7 300 47 From this data, determine this patient’s Km and Vmax for phenytoin.

Dose (mg/d) Css (mg/L) Dose Rate/Css (L/d) 100 3.7 27 300 47 6.4 JB is an 18 year male receiving phenytoin for prophylaxis of post-traumatic head injury seizures. The following steady state concentrations were obtained at the indicated doses: Dose (mg/d) Css (mg/L) Dose Rate/Css (L/d) 100 3.7 27 300 47 6.4 Vmax = 362 mg/d K0 (mg/d) Km = 9.7 mg/L K0/Css (L/d) April 7, 2019

Significance of Km Km is a constant Small Km means tight binding; high Km means weak binding Useful to compare Km for different substrates for one enzyme Hexokinase : D-fructose – 1.5 mM D-glucose – 0.15 mM Useful to compare Km for a common substrate used by several enzymes Hexokinase: D-glucose – 0.15 mM Glucokinase: D-glucose – 20 mM

Other applications 1)The Catalytic Efficiency kcat, the turnover number, is the number of substrate molecules converted to product per enzyme molecule per unit of time, when E is saturated with substrate. kcat/Km is an apparent second-order rate constant which measures how the enzyme performs when S is low

Competitive inhibitor 2) Vmax unaltered, Km increased

Noncompetitive inhibitor 3) Km unaltered, Vmax decreased

References Leon Shargel, A Text book of Applied Bio pharmaceutics and pharmacokinetics,5th edition Milo Gibaldi , Donald Perrier, Pharmacokinetics,2nd edition V.Venkateshwarlu, Biopharmaceutics and pharmacokinetics,(2004) Garret & Grisham , Biochemistry, 2nd edition http://en.wikipedia.org/wiki/Michaelis-Menten_kinetics

German biochemist and physician Canadian medical scientist

Thank You