Ohta Shinya, Tsuchida Ken, Choi Sunju, Sekine Yasuhiko, Shiga Yasuyuki, Ohtsubo Eiichi
Institute of Molecular and Cellular Biosciences, University of Tokyo, Bunkyo-ku, Japan.
J Bacteriol. 2002 Nov;184(22):6146-54. doi: 10.1128/JB.184.22.6146-6154.2002.
Transposases encoded by various transposable DNA elements and retroviral integrases belong to a family of proteins with three conserved acidic amino acids, D, D, and E, constituting the D-D-E motif that represents the active center of the proteins. IS1, one of the smallest transposable elements in bacteria, encodes a transposase which has been thought not to belong to the family of proteins with the D-D-E motif. In this study, we found several IS1 family elements that were widely distributed not only in eubacteria but also in archaebacteria. The alignment of the transposase amino acid sequences from these IS1 family elements showed that out of 14 acidic amino acids present in IS1 transposase, three (D, D, and E) were conserved in corresponding positions in the transposases encoded by all the elements. Comparison of the IS1 transposase with other proteins with the D-D-E motif revealed that the polypeptide segments surrounding each of the three acidic amino acids were similar. Furthermore, the deduced secondary structures of the transposases encoded by IS1 family elements were similar to one another and to those of proteins with the D-D-E motif. These results strongly suggest that IS1 transposase has the D-D-E motif and thus belongs to the family of proteins with the D-D-E motif. In fact, mutant IS1 transposases with an amino acid substitution for each of the three acidic amino acids possibly constituting the D-D-E motif were not able to promote transposition of IS1, supporting this hypothesis. The D-D-E motif identified in IS1 transposase differs from those in the other proteins in that the polypeptide segment between the second D and third E in IS1 transposase is the shortest, 24 amino acids in length. Because of this difference, the presence of the D-D-E motif in IS1 transposase has not been discovered for some time.
由各种转座DNA元件编码的转座酶和逆转录病毒整合酶属于一类蛋白质家族，该家族有三个保守的酸性氨基酸D、D和E，构成了代表蛋白质活性中心的D-D-E基序。IS1是细菌中最小的转座元件之一，它编码一种转座酶，一直被认为不属于具有D-D-E基序的蛋白质家族。在本研究中，我们发现了几种IS1家族元件，它们不仅广泛分布于真细菌中，也分布于古细菌中。对这些IS1家族元件的转座酶氨基酸序列进行比对后发现，在IS1转座酶中存在的14个酸性氨基酸中，有三个（D、D和E）在所有元件编码的转座酶的相应位置上是保守的。将IS1转座酶与其他具有D-D-E基序的蛋白质进行比较后发现，围绕这三个酸性氨基酸的多肽片段是相似的。此外，IS1家族元件编码的转座酶的推导二级结构彼此相似，也与具有D-D-E基序的蛋白质的二级结构相似。这些结果有力地表明，IS1转座酶具有D-D-E基序，因此属于具有D-D-E基序的蛋白质家族。事实上，对可能构成D-D-E基序的三个酸性氨基酸中的每一个进行氨基酸替换的突变IS1转座酶都无法促进IS1的转座，这支持了这一假说。在IS1转座酶中鉴定出的D-D-E基序与其他蛋白质中的不同之处在于，IS1转座酶中第二个D和第三个E之间的多肽片段最短，长度为24个氨基酸。由于这种差异，IS1转座酶中D-D-E基序的存在在一段时间内一直未被发现。
