Structural role of disulfide bridges in the cyclic ADP-ribose related bifunctional ectoenzyme CD38.
Human CD38, a type II cell surface glycoprotein, is a bifunctional ectoenzyme catalyzing both ADP-ribosyl cyclase and cyclic ADP-ribose (cADPR) hydrolase reactions. It shares a high degree of sequence homology with the cyclase from Aplysia species and studies of site-directed mutagenesis have recently demonstrated the importance, but not elucidated the role, of several cysteine residues highly conserved between these proteins. N-Ethylmaleimide, iodoacetamide and thiol-oxidizing reagents failed to affect either the cyclase or the weaker hydrolase activity of the Aplysia californica protein. Likewise, these reagents did not impair the two activities of CD38 purified from human erythrocytes. beta-mercaptoethanol had no effect on the Aplysia enzyme activities, while it inactivated both the cyclase and the cADPR hydrolase of CD38 by inducing its extensive oligomerization. In intact erythrocytes the beta-mercaptoethanol-dependent enzyme inactivation was completely prevented by prior cross-linking of the membrane proteins with glutaraldehyde. These data demonstrate that none of the cysteine residues plays any direct catalytic role in CD38 and Aplysia proteins, and that disulfide bridges are essential for maintaining the monomeric, catalytically active structure of CD38.
Guida
L
L
Institute of Biochemistry, University of Genoa, Italy.
Franco
L
L
Zocchi
E
E
De Flora
A
A
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000737:chemistry; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry*; D000943:Antigens, Differentiation / Q000737:chemistry*; D001048:Aplysia; D036563:Cyclic ADP-Ribose; D004220:Disulfides / Q000737:chemistry*; D006801:Humans; D008562:Membrane Glycoproteins; D008623:Mercaptoethanol / Q000494:pharmacology; D009699:N-Glycosyl Hydrolases / Q000737:chemistry*
D015703:Antigens, CD; D000943:Antigens, Differentiation; D004220:Disulfides; D008562:Membrane Glycoproteins; D036563:Cyclic ADP-Ribose; D000246:Adenosine Diphosphate Ribose; D008623:Mercaptoethanol; D009699:N-Glycosyl Hydrolases; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1