Roles of tryptophan residues on the Rhizopus delemar lipase activity: Chemical modification in a water-olive oil emulsion

Abstract

1. 1. The molecular weight of the lipase (EC 3.1.1.3) purified from Rhizopus delemar was 41 300. The average hydrophobicity, calculated from the results of amino acid analysis, was high. This suggests that a hydrophobic region of the lipase may play an important role in the formation of the enzyme triglyceride complex (ES complex). 2. 2. The tryptophan residues of the lipase were modified by 2-hydroxy 5 nitrobenzyl bromide (soluble in olive oil) and/or dimethyl-(2-hydroxy-5-nitrobenzyl) bromide (soluble in water) using the water-olive oil emulsion. It was found that one residue, modified from the olive oil side at the interface of the emulsion, was essential to the lipase activity and another one, modified from either the water or olive oil side might be a binding site. These two residues were buried in the lipase molecule unless the substrate was added, and were exposed to the enzyme surface when the enzyme-substrate complex was formed. Some speculation about the states of the other six residues is also presented.