Ca²⁺ Activation Kinetics of the Two Aspartate-Glutamate Mitochondrial Carriers, Aralar and Citrin: ROLE IN THE HEART MALATE-ASPARTATE NADH SHUTTLE

• Published in: The Journal of biological chemistry Vol. 282; no. 10; pp. 7098 - 7106
• Main Authors: Contreras, Laura, Gomez-Puertas, Paulino, Iijima, Mikio, Kobayashi, Keiko, Saheki, Takeyori, Satrústegui, Jorgina
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 09-03-2007
• Subjects: Amino Acid Sequence; Animals; Aspartic Acid - metabolism; Brain - metabolism; Calcium - metabolism; Glutamic Acid - metabolism; Kinetics; Malates - metabolism; Membrane Transport Proteins - chemistry; Membrane Transport Proteins - physiology; Mice; Mice, Inbred C57BL; Mitochondria, Heart - metabolism; Mitochondria, Liver - metabolism; Mitochondrial Membrane Transport Proteins; Mitochondrial Proteins - chemistry; Mitochondrial Proteins - physiology; Molecular Sequence Data; Muscle, Skeletal - metabolism; NAD - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M610491200

Ca²⁺ regulation of the Ca²⁺ binding mitochondrial carriers for aspartate/glutamate (AGCs) is provided by their N-terminal extensions, which face the intermembrane space. The two mammalian AGCs, aralar and citrin, are members of the malate-aspartate NADH shuttle. We report that their N-terminal extensions contain up to four pairs of EF-hand motifs plus a single vestigial EF-hand, and have no known homolog. Aralar and citrin contain one fully canonical EF-hand pair and aralar two additional half-pairs, in which a single EF-hand is predicted to bind Ca²⁺. Shuttle activity in brain or skeletal muscle mitochondria, which contain aralar as the major AGC, is activated by Ca²⁺ with S₀.₅ values of 280-350 nM; higher than those obtained in liver mitochondria (100-150 nM) that contain citrin as the major AGC. We have used aralar- and citrin-deficient mice to study the role of the two isoforms in heart, which expresses both AGCs. The S₀.₅ for Ca²⁺ activation of the shuttle in heart mitochondria is about 300 nM, and it remains essentially unchanged in citrin-deficient mice, although it undergoes a drastic reduction to about 100 nM in aralar-deficient mice. Therefore, aralar and citrin, when expressed as single isoforms in heart, confer differences in Ca²⁺ activation of shuttle activity, probably associated with their structural differences. In addition, the results reveal that the two AGCs fully account for shuttle activity in mouse heart mitochondria and that no other glutamate transporter can replace the AGCs in this pathway.