A new member of the leucyl aminopeptidase family purified and identified from a marine unicellular algae.

Leucyl aminopeptidase (LAP; EC 3.4.11.1) activity was purified from crude extracts of the marine unicellular algae Gonyaulax polyedra by a combination of hydrophobic interaction with phenyl sepharose, DEAE-cellulose, and mono-Q HR5/5 ion-exchange chromatography. The undenaturated protein has a molecular mass of about 110 kD and based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the enzyme appears to be composed of two possibly identical subunits of 55 kD. The identity of the protein was confirmed by a cross-reaction of the purified protein with an antibody raised against a commercial LAP. Biochemical characterization showed that the Gonyaulax enzyme was similar to most of the previously described LAPs. Gonyaulax LAP is a metallo-enzyme since EDTA and 1,10-phenathroline significantly inhibited activity. Addition of the metal ions Zn(2+), Cu(2+) inhibited 80% of LAP activity, suggesting they are not the natural cofactors of the enzyme. Other metals, such as Ca(2+), Co(2+), Mn(2+), or Mg(2+) (concentrations up to 4 mM), caused no alteration in the total activity of Gonyaulax LAP. Copyright 1999 Academic Press.