In previous papers we have reported that an aminopeptidase A (EC 3.4.11.7) purified from Staphylococcus chromogenes was able to catalyse the introduction of L-malic acid at the N-terminus of Tyr and Phe derivatives. We now show that this enzyme can be used for selective alpha-amino protection of derivatives of probably all amino acids, except Gly and Pro, by the malyl group. The following L-malyl derivatives were synthesized in thermodynamically controlled reactions with yields ranging from 4 to 47%: L-malyl-Tyr-OEt, -ALA-OMe, -Ser-OEt, -Lys-OEt, -Phe-OMe, -Met-NH2, -Glu-MH2, Arg-NH2, -Tye-NH2, -Val-NH2, -Ala-Phe and -Ala-Phe-NH2 (OEt and OMe are ethyl and methyl esters respectively). The reactions were monitored by reverse-phase h.p.l.c.; the products were quantified by amino acid analysis, and their structure was confirmed by m.s. No synthesis was obtained with Gly and Pro derivatives as nucleophiles. The effects of pH, temperature, enzyme concentration, nucleophile concentration, reaction time and addition of an organic co-solvent were studied. An important shift towards synthesis was obtained by carrying out the reactions at 55 degrees C in the presence of 55% organic co-solvent Triglyme (2,4,8,11-tetraoxadodecane) [8-60-fold increase in Ksyn. ([product] [acyl-donor]-1 [nucleophile]-1)].