The C-terminus of human bleomycin hydrolase is required for protection against bleomycin-induced chromosomal damage.
Mammalian bleomycin hydrolases (BH) are enzymes with proven exopeptidase activity responsible for deamidation of the beta-aminoalanine moiety in bleomycin and are thought to limit the therapeutic efficacy of the drug. We have recently determined that the highly conserved BH-like domain in the C-terminus of human bleomycin hydrolase (hBH) is critical both for in vitro aminopeptidase and bleomycin metabolizing activities. To determine if hBH protects mammalian cells against bleomycin clastogenic effect, we transfected CHO cells with plasmids encoding hBH or C-terminal truncated forms and evaluated the level of chromatid breaks after bleomycin exposure. CHO cells expressing hBH had 50% less chromatid breaks after bleomycin treatment compared with mock transfected cells. The eight amino acid bleomycin hydrolase-like domain in the C-terminus, which does not contain any of the putative active site amino acids, was essential for protection against bleomycin induced chromatid breaks. These results demonstrate that intracellular hBH levels can influence the clastogenic action of bleomycin and that the C-terminus has a functional role in the biological activity of hBH.
Lefterov
I M
IM
Department of Pharmacology and the Molecular Therapeutics/Drug Discovery Program of the University of Pittsburgh Cancer Institute, University of Pittsburgh, Biomedical Science Tower E1340, Pittsburgh, PA 15261, USA.
Koldamova
R P
RP
King
J
J
Lazo
J S
JS
D000596:Amino Acids; D000818:Animals; D001761:Bleomycin / Q000378:metabolism* / Q000494:pharmacology; D016466:CHO Cells; D019457:Chromosome Breakage; D006224:Cricetinae; D003546:Cysteine Endopeptidases / Q000737:chemistry* / Q000235:genetics; D016195:G2 Phase; D006801:Humans; D009153:Mutagens / Q000378:metabolism* / Q000494:pharmacology; D017384:Sequence Deletion
D000596:Amino Acids; D009153:Mutagens; D001761:Bleomycin; D003546:Cysteine Endopeptidases; C031262:bleomycin hydrolase