Nickel C M, Vandekerckhove J, Beyer P, Tadros M H
Institute for Biology II/Microbiology, University of Freiburg, Germany.
Gene. 1997 Jun 19;192(2):251-9. doi: 10.1016/s0378-1119(97)00085-1.
In Rhodobacter capsulatus (Rbc), the participation of DnaK in the synthesis of light harvesting antenna complex I (LHI) has been recently inferred from the finding that the amount of LHI alpha- and beta-polypeptides synthesized in an in vitro translation system was strongly reduced when DnaK was depleted. In the present work, a DnaK protein was isolated from Rbc and biochemically characterized. The N-terminus of the protein was sequenced and a corresponding oligo was used as probe in order to clone the gene coding for DnaK. The dnaK gene was located in an operon (dnaKJ) with two open reading frames, which code for DnaK and DnaJ, respectively. A promoter element corresponding to the consensus sequence of the atypical heat shock (HS) promoter of several alpha-purple proteobacteria was identified. Northern blot analysis indicated that dnaK and dnaJ belong to the same transcriptional unit; there were two transcripts, one comprising both the dnaK and dnaJ genes and a second with only dnaK. Primer extension analysis revealed that under both chemotrophic and phototrophic conditions transcription was initiated from the same position before and after HS. The promoter activity was studied under different growth conditions with a dnaK-lacZ fusion under the control of the dnaKJ promoter. The present work opens up the possibility to study the specific role of DnaK in the assembly of photosynthetic apparatus proteins.
在荚膜红细菌（Rbc）中，最近通过以下发现推断出DnaK参与了光捕获天线复合体I（LHI）的合成：当DnaK缺失时，体外翻译系统中合成的LHIα和β多肽的量大幅减少。在本研究中，从Rbc中分离出DnaK蛋白并进行了生化特性分析。对该蛋白的N端进行了测序，并使用相应的寡核苷酸作为探针来克隆编码DnaK的基因。dnaK基因位于一个操纵子（dnaKJ）中，该操纵子有两个开放阅读框，分别编码DnaK和DnaJ。鉴定出了一个与几种α-紫色变形菌的非典型热休克（HS）启动子的共有序列相对应的启动子元件。Northern印迹分析表明，dnaK和dnaJ属于同一个转录单元；有两种转录本，一种包含dnaK和dnaJ基因，另一种仅包含dnaK。引物延伸分析表明，在化学营养和光营养条件下，转录均从HS前后的同一位置起始。利用在dnaKJ启动子控制下的dnaK-lacZ融合体，研究了不同生长条件下的启动子活性。本研究为研究DnaK在光合装置蛋白组装中的特定作用开辟了可能性。
