Humboldt State University ® Department of Chemistry

Richard A. Paselk

Chem 431	Biochemistry	Fall 2008
Lecture Notes: 8 October	© R. Paselk 2008
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Enzyme Kinetics and Inhibition, cont.

Last time we left off our discussion with Competitive inhibition. next we have:

Noncompetitive inhibition

In Noncompetitive inhibition the inhibitor can bind to either E or ES. S & I do not bind to the same sites! (text Figure 6-15c)

Model: ; and .

Note that will have two inhibitor binding constants, they may be the same, as in the equation above, or could be different, leading to more complex behavior.

Plots for classic, simple situation (overhead MvH 11.5):

Mixed inhibition

Like non-competitive above,

Model: ; and

but with different values of K_i. This is in fact the more common situation, with a L-B plot with an intersection on the plot above the 1/v_o axis. (text Box 6-2 figure 3)

Uncompetitive inhibition

In uncompetitive inhibition the inhibitor binds ONLY to the ES complex (text Figure 6-15b).

Model: ; and .

For double reciprocal plots get parallel lines! (text Box 6-2 figure 2) This is not generally found for single substrate enzymes, but is found in multi-substrate systems.

Multi-substrate Enzyme Kinetics

Look at three common and easily understood types. We will use Cleland Nomenclature and "Kinetic mechanism diagrams."

• Ordered Sequential Bi Bi mechanism (two on; two off); Note: A must bind first, Q is released last.
• Ping Pong Bi Bi (one on, one off; one on, one off); Note: have some sort of modified enzyme intermediate (often covalent intermediate)
• Random Sequential Bi Bi (two on; two off); Note: A or B may bind first, P or Q may be released last.

Note that in each case we can predict/explain the pattern of inhibition on the basis of the substrate and inhibitor binding to the same "enzyme form." Thus for the Ordered Sequential mechanism only the first substrate and last product bind to the same form, in this case the free enzyme. Similarly for the Ping pong mechanism the first substrate and last product should be competitive as the both bind the free enzyme. In this case we also see a competitive inhibition between the second substrate and the first product, since they both bind to the E-X complex. The Random Sequential mechanism is a bit more subtle. Here we see across the board noncompetitive since in each case the substrates (and products) can each bind to more than one substrate form, so competitive inhibition will not be possible! (Think of the product as competing with one order of binding but not the other.)

Temperature Effects on Enzymes (and Proteins)

Temperature profile reflects two underlying phenomena:

• The effects of temperature on rate due to the fact that as temp. increases more molecules will have the energy required to achieve activation energy, E_a. (Review: Arrhenius equation: , and the Maxwell-Boltzman Distribution.)
• The effects of temperature on protein stability: at high temperatures the protein denatures.

Together these effects lead to the plot below where the rising leg is due to activation energy effects (increasing rate) and the falling leg is due to protein denaturation.

Pathway Diagrams

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Last modified 8 October 2008