Essential cysteine residues for cyclic ADP-ribose synthesis and hydrolysis by CD38.
We have recently demonstrated that cyclic ADP-ribose (cADPR) serves as a second messenger for glucose-induced insulin secretion (Takasawa, S., Nata, K., Yonekura, H., and Okamoto, H. (1993) Science 259, 370-373) and that human leukocyte antigen CD38 has both ADP-ribosyl cyclase and cADPR hydrolase activities (Takasawa, S., Tohgo, A., Noguchi, N., Koguma, T., Nata, K., Sugimoto, T., Yonekura, H., and Okamoto, H. (1993) J. Biol. Chem. 268, 26052-26054). Although the amino acid sequence of Aplysia ADP-ribosyl cyclase exhibits a high degree of amino acid sequence identity with that of CD38, the Aplysia enzyme shows only ADP-ribosyl cyclase but not cADPR hydrolase. In the present study, we introduced site-directed mutations to CD38 and found that C119K- and/or C201E-CD38 exhibited only ADP-ribosyl cyclase activity. Furthermore, Aplysia ADP-ribosyl cyclase into which we introduced the mutations K95C and E176C, which correspond to residues 119 and 201 of human CD38, exhibited not only ADP-ribosyl cyclase activity but also cADPR hydrolase. These results indicate that cysteine residues 119 and 201 in CD38 have crucial roles in the synthesis and hydrolysis of cADPR.
Tohgo
A
A
Department of Biochemistry, Tohoku University School of Medicine, Miyagi, Japan.
Takasawa
S
S
Noguchi
N
N
Koguma
T
T
Nata
K
K
Sugimoto
T
T
Furuya
Y
Y
Yonekura
H
H
Okamoto
H
H
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D000943:Antigens, Differentiation / Q000378:metabolism*; D001048:Aplysia; D001483:Base Sequence; D036563:Cyclic ADP-Ribose; D003545:Cysteine / Q000378:metabolism*; D018076:DNA, Complementary; D006801:Humans; D006868:Hydrolysis; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016415:Sequence Alignment
D015703:Antigens, CD; D000943:Antigens, Differentiation; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D036563:Cyclic ADP-Ribose; D000246:Adenosine Diphosphate Ribose; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1; D003545:Cysteine