Roles of adjoining Asp and Cys residues of first matrix-facing loop in transport activity of yeast and bovine mitochondrial ADP/ATP carriers.
The mitochondrial ADP/ATP carrier (AAC) transports substrate by interconversion of its conformation between m- and c-states. The 1st loop facing the matrix (LM1) is extruded into the matrix in the m-state and is suggested to intrude into the mitochondrial membrane on conversion to the c-state conformation [Hashimoto, M., Majima, E., Goto, S., Shinohara, Y., and Terada, H. (1999) Biochemistry 38, 1050-1056]. To elucidate the mechanism of the translocation of LM1, we examined the effects of site-directed mutagenesis of two adjoining residues, Cys56 and Asp55 in the bovine type 1 AAC and Cys73 and Asp72 in the yeast type 2 AAC, on the substrate transport activity. We found that (i) replacement of the Cys by bulky and hydrophilic residues was unfavorable for efficient transport activity, (ii) the carboxyl groups of the Asp residues of the bovine and yeast AACs were essential and strictly position-specific, and (iii) hence, the mutation to Glu showed transport activity comparable to that of the native AACs. Based on these results, we discussed the functional role of LM1 in the transport activity of AAC.
Faculty of Pharmaceutical Sciences, University of Toyama, 2630, Sugitani, Toyama 930-0194.
Yoshitaka
Y
Kihira
Mitsuru
M
Hashimoto
Yasuo
Y
Shinohara
Eiji
E
Majima
Hiroshi
H
Terada
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000235:genetics* / Q000378:metabolism; D002417:Cattle; D003545:Cysteine / Q000235:genetics* / Q000378:metabolism; D051336:Mitochondrial Membranes / Q000378:metabolism*; D008969:Molecular Sequence Data; D033704:Nucleotide Transport Proteins / Q000235:genetics* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D033704:Nucleotide Transport Proteins; D029701:Saccharomyces cerevisiae Proteins; D001224:Aspartic Acid; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D003545:Cysteine