Mg2+-dependent interactions of ATP with the cystathionine-β-synthase (CBS) domains of a magnesium transporter.
Ancient conserved domain protein/cyclin M (CNNM) family proteins are evolutionarily conserved Mg(2+) transporters. However, their biochemical mechanism of action remains unknown. Here, we show the functional importance of the commonly conserved cystathionine-β-synthase (CBS) domains and reveal their unique binding ability to ATP. Deletion mutants of CNNM2 and CNNM4, lacking the CBS domains, are unable to promote Mg(2+) efflux. Furthermore, the substitution of one amino acid residue in the CBS domains of CNNM2, which is associated with human hereditary hypomagnesemia, abrogates Mg(2+) efflux. Binding analyses reveal that the CBS domains of CNNM2 bind directly to ATP and not AMP in a manner dependent on the presence of Mg(2+), which is inhibited in a similar pattern by the disease-associated amino acid substitution. The requirement of Mg(2+) for these interactions is a unique feature among CBS domains, which can be explained by the presence of highly electronegative surface potentials around the ATP binding site on CNNM2. These results demonstrate that the CBS domains play essential roles in Mg(2+) efflux, probably through interactions with ATP. Interactions with ATP, which mostly forms complexes with Mg(2+) in cells, may account for the rapid Mg(2+) transport by CNNM family proteins.
From the Department of Cellular Regulation, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan and.
Yusuke
Y
Hirata
From the Department of Cellular Regulation, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan and.
Yosuke
Y
Funato
the Laboratory of Protein Informatics, Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
Yu
Y
Takano
From the Department of Cellular Regulation, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan and hmiki@biken.osaka-u.ac.jp.
Hiroaki
H
Miki
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D027682:Cation Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003541:Cystathionine beta-Synthase / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D017136:Ion Transport / Q000235:genetics; D008274:Magnesium / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity
C539261:CNNM4 protein, human; D027682:Cation Transport Proteins; C510868:Cnnm2 protein, mouse; D000255:Adenosine Triphosphate; D003541:Cystathionine beta-Synthase; D008274:Magnesium
AMP; ATP; Ancient Conserved Domain Protein/Cyclin M (CNNM); Cystathionine-β-synthase (CBS) Domain; Magnesium; Magnesium Homeostasis; Magnesium Transport; Transporters