Kim S K, Shim J
Department of Molecular and Cellular Biology, Harvard University, 7 Divinity Avenue, Cambridge, Massachusetts 02138, USA.
Biochem Biophys Res Commun. 1999 Sep 16;263(1):113-7. doi: 10.1006/bbrc.1999.1317.
Two different yeast two hybrid systems were used to examine interaction between the partition proteins SopA and SopB of F plasmid as well as their self association. In one system, the yeast Gal4 protein DNA binding domain (Gal4-BD) is fused to the N-terminus of the bait protein, and the Gal4 activation domain (Gal4-AD) is fused to the N-terminus of the target protein (1). In the other system, the target hybrid remains unchanged but E. coli LexA protein (LexA) is fused to the C-terminus of the bait protein (2). It is found that C-terminus part of SopB is involved in interaction with itself, as an N-terminal truncation of SopB, SopB-(120-323) remains capable of self association. For interaction between SopA, deletion of the N-terminal part weakens but does not abolish the interaction. Interaction between SopB and SopA protein was also detected, but only by the use of the second system. Full length SopB [SopB-(1-323)] or SopB-(1-180) lacking the C-terminal region beyond amino acid 180 can interact with full-length SopA-(1-383) protein.
使用两种不同的酵母双杂交系统来检测F质粒的分配蛋白SopA和SopB之间的相互作用以及它们的自我缔合。在一种系统中，酵母Gal4蛋白DNA结合结构域（Gal4-BD）与诱饵蛋白的N端融合，Gal4激活结构域（Gal4-AD）与靶蛋白的N端融合（1）。在另一种系统中，靶杂交体保持不变，但大肠杆菌LexA蛋白（LexA）与诱饵蛋白的C端融合（2）。发现SopB的C端部分参与其自身相互作用，作为SopB的N端截短形式，SopB-(120 - 323)仍然能够自我缔合。对于SopA之间的相互作用，N端部分的缺失会减弱但不会消除这种相互作用。还检测到SopB和SopA蛋白之间的相互作用，但仅通过使用第二种系统检测到。全长SopB [SopB-(1 - 323)]或缺少180位氨基酸之后C端区域的SopB-(1 - 180)可以与全长SopA-(1 - 383)蛋白相互作用。
