This invention describes a method of removing N-terminal alanine residues
from polypeptides, preferably recombinant proteins, using an
aminopeptidase derived from the marine bacterium Aeromonas proteolytica.
Accordingly, Aeromonas aminopeptidase (AAP; E.C. 3.4.11.10) can be used to
remove N-terminal alanyl residues from derivatives of human somatotropin
(hST, human growth hormone, or hGH), porcine somatotropin (pST), and
bovine somtotropin (bST), for example, to yield proteins having their
native amino acid sequences. The enzyme reactions can be carried out in
free solution, or the AAP can be immobilized on a solid support, for
reactions carried out in vitro. An efficient method for converting Ala-hGH
to hGH, for example, comprises expression of Ala-hGH in E. coli, recovery
of inclusion bodies, solubilization and refolding in detergent, detergent
removal by ultrafiltration, selective precipitation, enzyme cleavage,
followed by two column chromatography steps.
