Department of Pathology, College of Medicine, University of Florida, Gainesville, Florida, United States of America.
PLoS One. 2011 Apr 21;6(4):e19250. doi: 10.1371/journal.pone.0019250.
The adenine nucleotide translocase (ANT) mediates the exchange of ADP and ATP across the inner mitochondrial membrane. The human genome encodes multiple ANT isoforms that are expressed in a tissue-specific manner. Recently a novel germ cell-specific member of the ANT family, ANT4 (SLC25A31) was identified. Although it is known that targeted depletion of ANT4 in mice resulted in male infertility, the functional biochemical differences between ANT4 and other somatic ANT isoforms remain undetermined. To gain insight into ANT4, we expressed human ANT4 (hANT4) in yeast mitochondria. Unlike the somatic ANT proteins, expression of hANT4 failed to complement an AAC-deficient yeast strain for growth on media requiring mitochondrial respiration. Moreover, overexpression of hANT4 from a multi-copy plasmid interfered with optimal yeast growth. However, mutation of specific amino acids of hANT4 improved yeast mitochondrial expression and supported growth of the AAC-deficient yeast on non-fermentable carbon sources. The mutations affected amino acids predicted to interact with phospholipids, suggesting the importance of lipid interactions for function of this protein. Each mutant hANT4 and the somatic hANTs exhibited similar ADP/ATP exchange kinetics. These data define common and distinct biochemical characteristics of ANT4 in comparison to ANT1, 2 and 3 providing a basis for study of its unique adaptation to germ cells.
腺嘌呤核苷酸转位酶（ANT）介导 ADP 和 ATP 在线粒体内膜之间的交换。人类基因组编码多种以组织特异性方式表达的 ANT 同工型。最近，鉴定出了一种新的、生殖细胞特异性的 ANT 家族成员 ANT4（SLC25A31）。尽管已知在小鼠中靶向敲除 ANT4 会导致雄性不育，但 ANT4 与其他体细胞 ANT 同工型之间的功能生化差异仍未确定。为了深入了解 ANT4，我们在酵母线粒体中表达了人源 ANT4（hANT4）。与体细胞 ANT 蛋白不同，hANT4 的表达未能补充 AAC 缺陷酵母菌株在需要线粒体呼吸的培养基上的生长。此外，来自多拷贝质粒的 hANT4 的过表达会干扰酵母的最佳生长。然而，hANT4 的特定氨基酸突变改善了酵母线粒体的表达，并支持 AAC 缺陷酵母在不可发酵碳源上的生长。这些突变影响了与磷脂相互作用的预测氨基酸，表明脂质相互作用对该蛋白功能的重要性。每个突变的 hANT4 和体细胞 hANTs 都表现出相似的 ADP/ATP 交换动力学。这些数据定义了 ANT4 与 ANT1、2 和 3 相比的常见和独特的生化特征，为研究其对生殖细胞的独特适应提供了基础。
