Purification and preliminary characterization of dDAP, a novel dipeptidylaminopeptidase from Dictyostelium discoideum.
We have discovered and purified a novel dipeptidylaminopeptidase (DAP) from the cell-free broth of Dictyostelium discoideum Ax3. The enzyme is secreted in parallel with cell growth in axenic broth culture. It shares substrate preferences with both DAP-I and DAP-III enzymes yet is distinct from both in some physical properties. Similar to DAP-I, the D. discoideum enzyme is able to cleave a variety of dipeptides from the amino termini of substrates. In addition, it readily cleaves substrate sequences beginning with RR- and KK-, a property of the DAP-III class. The D. discoideum enzyme has a pH optimum of 3.5, a subunit molecular mass of 66,000 daltons, and a molecular weight of approximately 225,000 and is not significantly inhibited by cysteine or serine protease inhibitors. It is inhibited by leupeptin and trivalent cations. On the basis of enzymological and other data presented here, we conclude the D. discoideum enzyme does not belong to any of the previously reported DAP classes (DAP-I, -II, -III, -IV) and propose that the class DAP-V be established with this D. discoideum enzyme as the first member.
Atkinson
P R
PR
Fermentation Process Development, Lilly Research Laboratories, Eli Lilly & Company, Indianapolis, Indiana 46285, USA.
Hilton
M D
MD
Lambooy
P K
PK
D000595:Amino Acid Sequence; D000818:Animals; D020789:Cathepsin C; D002852:Chromatography, Ion Exchange; D004023:Dictyostelium / Q000201:enzymology*; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005285:Fermentation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity
D010455:Peptides; D011480:Protease Inhibitors; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D020789:Cathepsin C; C100152:dipeptidyl peptidase III