Leveson-Gower Dennis B, Michnick Stephen W, Ling Victor
British Columbia Cancer Research Centre, British Columbia Cancer Agency, University of British Columbia, Vancouver, V5Z 1L3 Canada.
Biochemistry. 2004 Nov 9;43(44):14257-64. doi: 10.1021/bi0491245.
The transporter associated with antigen presentation (TAP) is an ATP-binding cassette (ABC) protein which transports peptides for presentation to the immune system. TAP is composed of two half transporters, TAP1 (ABCB2) and TAP2 (ABCB3), which heterodimerize to function. In humans, the TAP family consists of TAP1, TAP2, and TAPL (ABCB9). While the TAP1-TAP2 complex is well characterized, TAPL's dimerization state and function are unknown. To identify interactions within the human TAP family, we adapted the dihydrofolate reductase protein-fragment complementation assay (DHFR PCA) to half ABC transporters. This assay has been shown to be suitable for the study of membrane-bound proteins in vivo [Remy, I., Wilson, I. A., and Michnick, S. W. (1999) Science 283, 990-993]. With this method, in vivo TAP1-TAP2 heterodimerization was confirmed, no homodimerizations were detected with TAP1 or TAP2, and TAPL did not show any interaction with TAP1 or TAP2. However, we found strong evidence that TAPL forms homodimers. These results provide evidence of a novel homomeric TAPL interaction and demonstrate that the DHFR PCA will be of general utility in studies of half ABC transporter interactions in vivo.
抗原呈递相关转运体（TAP）是一种ATP结合盒（ABC）蛋白，负责转运肽段以呈递给免疫系统。TAP由两个半转运体TAP1（ABCB2）和TAP2（ABCB3）组成，二者异源二聚化后发挥功能。在人类中，TAP家族包括TAP1、TAP2和TAPL（ABCB9）。虽然TAP1 - TAP2复合物已得到充分表征，但TAPL的二聚化状态和功能尚不清楚。为了确定人类TAP家族内部的相互作用，我们将二氢叶酸还原酶蛋白片段互补分析（DHFR PCA）应用于半ABC转运体。该分析已被证明适用于体内膜结合蛋白的研究[雷米，I.，威尔逊，I. A.，和米奇尼克，S. W.（1999年）《科学》283，990 - 993]。通过这种方法，在体内证实了TAP1 - TAP2异源二聚化，未检测到TAP1或TAP2的同二聚化，并且TAPL与TAP1或TAP2未显示任何相互作用。然而，我们发现有力证据表明TAPL形成同二聚体。这些结果为一种新型的TAPL同聚体相互作用提供了证据，并证明DHFR PCA在体内半ABC转运体相互作用研究中将具有普遍用途。
