(DEC 2013 GU)
43. The K_cat/K_m values of an enzyme-catalyzed reaction when plotted as a function of pH yielded a bell-shaped curve with a maximum around pH 6.0. Which of the following conclusions best describes the above observation?
(1) A His residue may be important for catalysis.
(2) The enzyme would be highly stable at pH 6.0.
(3) The enzyme would be most active below and above pH 6.0.
(4) The enzyme will require a metal ion as cofactor for catalysis.

The correct answer is (1) A His residue may be important for catalysis.

Introduction

The activity of enzymes is highly sensitive to pH changes, often displaying bell-shaped curves when plotting catalytic efficiency (kcat/Kmkcat/Km) versus pH. Such profiles suggest the involvement of ionizable amino acid side chains that must be in specific protonation states to facilitate catalysis effectively. This article examines the interpretation of a bell-shaped pH dependence curve peaking around pH 6.0, highlighting the probable role of histidine residues in the enzymatic reaction mechanism.

Understanding kcat/Kmkcat/Km and Its pH Dependence

• The ratio kcat/Kmkcat/Km is a measure of catalytic efficiency, combining substrate binding (affinity) and conversion rate.

• pH affects ionization states of enzyme residues and substrate, influencing both binding and catalysis.

• A bell-shaped curve indicates two ionizable groups must be in particular protonation states, often requiring one to be protonated and another deprotonated for optimal activity.

Why Histidine?

• Histidine side chain has a pKa around 6.0, well-aligned with the peak catalytic efficiency pH.

• It often acts as a proton donor or acceptor in enzyme active sites, critical for catalytic mechanisms such as general acid/base catalysis.

• The observed kcat/Kmkcat/Km bell curve around pH 6.0 strongly suggests involvement of a histidine residue essential for enzyme activity.

Other Options Explained

• (2) High enzyme stability at pH 6.0:
  Stability is different from catalysis; bell-shaped catalytic efficiency does not directly imply maximal stability.

• (3) Most active below and above pH 6.0:
  Bell shape shows maximal activity at pH 6.0, not outside it.

• (4) Requirement of metal ion cofactor:
  pH dependence alone does not indicate metal ion dependence, which would demand other experimental evidence.

Biological Significance

• Identifying ionizable residues involved in catalysis informs enzyme mechanism elucidation.

• Enables rational enzyme engineering or inhibitor design.

• Guides optimal pH settings for industrial and research applications.

Summary Table

Conclusion

A bell-shaped profile of catalytic efficiency (kcat/Kmkcat/Km) centered at pH 6.0 strongly supports that a histidine residue, with a side-chain pKa near this pH, plays a crucial role in the enzyme’s catalytic mechanism. This insight provides valuable guidance for enzymologists studying reaction mechanisms and those optimizing enzyme use.

This detailed interpretation assists students and researchers in connecting enzyme function with underlying chemical protonation dynamics in biological catalysis.