Biochemistry I Fall Term, 2004 Serine Proteases: pH Effects in Enzyme Mechanism In many enzymes, the ionization state of the amino acid side chains are important for catalytic activity. In the case of trypsin, the His residue in the active site must be deprotonated in order for it to act as a general base catalyst. Consider the following reaction scheme: Ka is the acidity constant for the group on the enzyme (i.e. His in the case of trypsin) The effect of the protonation on the enzyme activity can be calculated using steady-state methods. Note: The full derivation can found on pH Effects in Enzyme Mechanism, linked from the Lecture 26 notes. The result we will consider is a modified Michaelis-Menten equation, in which both Vmax and KM are modified: and where: Thus, a hyperbolic substrate saturation plot will be obtained at all pH values! Compare the above to the analogous expressions we found for enzyme inhibition. How Vmax varies with [H+] can be investigated by looking at some values for [H+]: [H+] >>Ka (low pH) = Ka <<Ka (high pH) large 2 1 V'max --> 0 Vmax/2 Vmax The above shows that when the pH is such that the velocity of the reaction is reduced by half, the pH is equal to the pKa of the group that is involved in catalysis. The above table is an accurate summary of the dependence of Vmax on pH for chymotrypsin in the range where the protein is in its native conformation. (Wha...