Lawson J E, Gawaz M, Klingenberg M, Douglas M G
Department of Biochemistry and Biophysics, University of North Carolina Medical School, Chapel Hill 27514.
J Biol Chem. 1990 Aug 25;265(24):14195-201.
The gene encoding the major ADP/ATP carrier in yeast AAC2 (pet9; Lawson, J., and Douglas, M. (1988) J. Biol. Chem. 263, 14812-14818) has been disrupted (delta AAC2) by itself and in combination with a disruption of a second translocator gene AAC1 (delta AAC1). Disruption of AAC2 like the pet9 mutation renders yeast unable to grow on a nonfermentable carbon source. The AAC1 AAC2 double disruption exhibits a phenotype identical to the AAC2. This provides the host strain for the analysis of point mutations in the AAC protein. We have initiated this structure-function analysis by characterizing and confirming that the pet9 mutation is a G to A transition resulting in an arginine to histidine change at position 96. Site-directed replacements at Arg96 confirm its essential function for growth on a nonfermentable carbon source. These data also suggest that in the absence of functional AAC1 and AAC2 gene products, adenine nucleotide transport across the mitochondrial inner membrane must occur by an as yet unidentified translocator or translocation mechanism or that within these cells separate intra- and extramitochondrial adenine nucleotide pools can exist to support growth.
酵母中主要ADP/ATP载体的编码基因AAC2（pet9；劳森，J.，和道格拉斯，M.（1988年）《生物化学杂志》263，14812 - 14818）已被单独破坏（ΔAAC2），并与第二个转运体基因AAC1的破坏（ΔAAC1）相结合。与pet9突变一样，AAC2的破坏使酵母无法在非发酵碳源上生长。AAC1 AAC2双破坏表现出与AAC2相同的表型。这为分析AAC蛋白中的点突变提供了宿主菌株。我们通过表征和确认pet9突变是一个从G到A的转变，导致第96位的精氨酸变为组氨酸，从而启动了这种结构 - 功能分析。在精氨酸96处的定点替换证实了其对于在非发酵碳源上生长的基本功能。这些数据还表明，在缺乏功能性AAC1和AAC2基因产物的情况下，腺嘌呤核苷酸穿过线粒体内膜的转运必定通过一种尚未确定的转运体或转运机制发生，或者在这些细胞内可以存在独立的线粒体内外腺嘌呤核苷酸池来支持生长。
