Mono-p-nitrophenyl fumarate

The polypeptide KO-42 catalyzed the hydrolysis of mono-p-nitrophenyl fumarate with a second-order rate constant ki of 0.1 M s , a rate enhancement of more... 

Figure 5.5. The 42-residue catalyst JNII has enhancement over that of the 4-two His residues that catalyze the hydrolysis of methylimidazole catalyzed reaction is the mono-p-nitrophenyl fumarate by a combination largest at a pH below/that of both His of nucleophilic and general-acid catalysis. The residues, p/fa values are indicated and the relative rate...

The Bronsted coefficient p for imidazole catalysis of p-nitrophenyl acetate hydrolysis is 0.8 [29], and the second-order rate constants of all His residues can therefore be related to that of 4-methylimidazole to determine whether there are effects on reactivity beyond those of differential nudeophilicity and levels of protonation. The reactivity of His residues in the pH independent region may be estimated from rate constants, pH and values. The second-order rate constant of the 4-methylimidazole catalyzed hydrolysis of mono-p-nitrophenyl fumarate at pH 5.85 and 290 K is 1.02 x 10 s. From this value and the pfC of 7.9, the second-... 

The nature of the cooperativity was further characterized based on results from kinetic measurements. The two HisH+-His pairs in helix II catalyzed the hydrolysis of mono-p-nitrophenyl fumarate at pH 5.1 and 290 K with second-order rate constants of 0.01 M-i s-i (JNI, His-26, His-30) and 0.055 M s (JNII, His-30, His-34), respectively, and a rate constant ratio JNII/JNI of 5.5. The pKa values of both His residues in JNII are the same, so for the analysis it does not matter which residue is the nucleophile and which one is the acid. In JNI, however, the pKa values are 6.9 for His-26 and 5.6 for His-30. The rate constant ratio of 5.5 should therefore arise due to the difference in nucleophihcity or due to the difference in acidity, or if both residues in the pair can be both nucleophile and acid, from a mixture of the two. If His-30 functions as a general acid in JNI, then the rate constant ratio should arise from the difference in nucleophUicity between two nucleophiles with the pKa values 5.6 and 6.9. We can, however, calculate the reactivity difference as in Section 5.2.3 to find that 10 - = 1.8, one third of the observed ratio of 5.5. If, on the other hand, the rate constant ratio is due to a difference in general-acid catalysis by two residues with pKa values of 5.6 and 6.9, then the the Bronsted equa-hon for general-acid catalysis can be applied... 

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