Effects on tryptophyl absorption of the ionization of the catalytic carboxyls in hen and turkey lysozymes

Area of Science:

• Biochemistry
• Enzymology
• Protein Spectroscopy

Background:

• Lysozymes are enzymes crucial for bacterial cell wall hydrolysis.
• Understanding the enzymatic mechanism requires knowledge of protein structure and protonation states.
• Tryptophyl residues play roles in enzyme structure and function.

Purpose of the Study:

• To compare the spectral properties of hen and turkey egg-white lysozymes.
• To determine the pK values of catalytic carboxyl groups (Glu 35 and Asp 52) in both lysozymes.
• To investigate the interaction between tryptophyl residues and catalytic groups.

Main Methods:

• Measurement of difference spectra of hen and turkey egg-white lysozymes upon acidification.
• Analysis of pH dependence of extinction differences at 295 nm and 301 nm.
• Comparison with circular dichroism data for validation.

Main Results:

• Both lysozymes showed spectral peaks at 295 nm and 301 nm, characteristic of tryptophyl residues.
• pH dependence curves for extinction differences were identical for both hen and turkey lysozymes.
• Determined macroscopic pK values for Glu 35 and Asp 52 were 6.0 and 3.3, respectively, consistent with previous circular dichroism studies.

Conclusions:

• Hen and turkey egg-white lysozymes share similar spectral characteristics and catalytic residue protonation states.
• The determined pK values provide insights into the active site environment of lysozymes.
• The spectral changes at 295 nm suggest factors beyond simple electrostatic effects influence tryptophyl residue behavior.