EFFECT OF BESTATIN ANALOGS AND OTHER COMPOUNDS ON ENKEPHALIN HYDROLYSIS BY AN AMINOPEPTIDASE FROM THE MESOPHILES PSEUDOMONAS-SP ATCC-11299A AND CHROMOBACTERIUM VIOLACEUM ATCC-12540

  • 1 January 1983
    • journal article
    • research article
    • Vol. 39  (3) , 463-475
Abstract
In studies on newly synthesized compounds for their potential analgesic effect, non-mammalian sources were investigated for the presence of enkephalin degrading enzymes. An aminopeptidase that catalyzes the hydrolysis of the tyrosyl-glycyl bond of Leu- and Met-enkephalin was purified from the mesophiles Pseudomonas sp. ATCC 11299a (Ps) and Chromobacterium violaceum ATCC 12540 (Cv). Each preparation hydrolyzed to varying extents neutral dipeptides, tripeptides, tetrapeptides and amino acid .beta.-naphthylamides. The Ps enzyme has a pH optimum of 6.8, Km of 80 .mu.M and a Vmax of 6.7 nmol/min per mg of protein. The Cv enzyme has a pH optimum of 6.8-7.2, Km of 111 .mu.M and a Vmax of 42 nmol/min per mg of protein. Both are sulfhydryl enzymes since they are activated by dithiothreitol (DTT) and inactivated by p-chloro- and p-hydroxymercuribenzoate. They are not glycoproteins since they pass unretained through a Con A-Sepharose column. The activity lost by dialysis against EDTA can be restored, totally or in part, by Co2+, Mg2+, Mn2+ and Ni2+ ions exerting an inhibitory effect were Al3+, Cd2+, Cu2+, Hg2+ and Zn2+. From a range of organic compounds, the greatest inhibition was elicited by the microbial peptides amastatin and bestatin. Several dipeptide analogs of bestatin, synthesized from DL-threo-2-amino-3-hydroxy-3-phenylpropanoic acid (AHPP) as the N-terminal residue to define the stereospecific requirements of the .alpha.,.beta.-functional groups for maximal activity, were not as active as the parent compound.