BCHM 307 1st Edition Lecture 12 Outline of Last Lecture I. Mass SpectrometryA. Definition of Ionization SourceB. Definition of Mass AnalyzerC. Definition of DetectorII. In-Gel Protease DigestionIII. Unknown Protein AnalysisA. Peptide Mass FingerprintingB. Tandem Mass SpectrometryIV. EnzymesA. Definition of EnzymeB. Definition of SubstrateV. The Two Enzyme ModelOutline of Current Lecture I. Enzyme ReactionII. Enzyme Rate EquationIII. Michaelis-Menten PlotA. Definition of Enzyme VelocityB. Velocity EquationC. Michaelis ConstantCurrent LectureThe way that enzymes interact with substrates can be shown by the following equation. E + S ⇋ ES ⇋ ES* ⇋ EP ⇋ E + PE is the enzyme, S is the substrate, ES is the enzyme/substrate complex, ES* is the enzyme transition state complex, EP is the enzyme product complex and P is the product. The place where the substrate binds is called the active site. The active site is like a pocket in the enzyme, which is made by protein folds. Substrates can use hydrogen bonding, hydrophobic interactions,and ionic interactions to promote binding to the active site. The various complexes mentioned above represent a mixture of enzyme and substrate. The ES* is neither substrate nor product, but a transition point. Once the product is formed, theenzyme will release it. The reaction can be simplified to A +B ⇋ P, where A and B are two These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.reactants and P is the product formed after the reaction has been catalyzed by an enzyme. The rate of the reaction is a function of change in time. It can be defined three different ways shownbelow, based on the decreasing amount of reactant or increasing amount of product.−∆ [ A ]∆ t −∆ [B]∆ t ∆[ P]∆ tThe quality of the enzyme activity, or assay, can be determined through various methods. It can be based on the disappearance of the substrate or appearance of the product. The change in the gain of a product/loss of a substrate over time is called the velocity. Velocity of a reaction that is catalyzed by an enzyme depends upon the concentration of the substrate. It should be noted that it is not over a change in time. This data can be plotted in what is known as a Michaelis-Menten plot. This graph will give you a hyperbolic shape. The initial velocity is plotted on the y-axis. The concentration of the substrate over time is plotted onthe x-axis. The velocity will increase as the substrate concentration increase, until the enzyme is fully saturated. The graph will level off at this point and the added substrate will not change the velocity of the reaction. During this whole process the amount of enzyme will not change.The Michaelis-Menton equation is V =Vmax x [ S]Km+[S ]. It is based on the simplified version of the enzyme reaction, where K1 and K3 are forward rates and K2 is the reverse rate.E + S ⇋ ES → E + P. This equation looks at the initial rates when [P] is close to 0.For the above velocity equation V is the velocity at a given substrate concentration, [S]. Vmax is the maximum velocity seen when the enzyme is fully saturated, called the zero order reaction. Km is called the Michaelis constant and can be found by the following:Km=K 2+K 3K 1 , which is approximately equal to[E][ S][ ES]. A low Km indicates a high affinityfor the
View Full Document
Unlocking...