Formylation-independent eubacterial protein initiation

引发剂甲硫基trna用于启动古细菌和真核生物细胞质中的蛋白质合成。相反，真细菌和某些真核细胞器（如线粒体和叶绿体）中的蛋白质合成可以使用引发剂甲酰基-蛋氨酸- trna （fMet-tRNAfMet）启动。甲酰基化对引发剂甲硫基trna （Met-tRNAfMet）具有特异性，并由甲硫基trna甲酰基转移酶（MTF）催化。体外和体内的早期研究使人们普遍认为Met-tRNAfMet的甲酰化是所有真细菌蛋白质合成起始的关键步骤。这一观点在大肠杆菌、肺炎链球菌和枯草芽孢杆菌中发现MTF基因的破坏严重限制了细胞生长，并且MTF基因存在于迄今测序的所有真细菌基因组中，这一发现加强了这一观点。然而，这种真细菌蛋白质起始的教条不再有效，因为一些细菌已被证明可以独立于甲酰化启动蛋白质合成。我的实验室首先证明了一种真细菌，铜绿假单胞菌，有能力利用Met-tRNAfMet来启动蛋白质合成。随后，研究表明，在一定的生长条件下，金黄色葡萄球菌、流感嗜血杆菌和酿酒酵母线粒体中，Met-tRNAfMet的甲酰化对于蛋白质的起始也不是必需的。因此，与大肠杆菌、肺炎链球菌和枯草芽孢杆菌依赖于fMet-tRNAfMet进行蛋白起始不同，铜绿假单胞菌、金黄色葡萄球菌和流感嗜血杆菌可以使用fMet-tRNAfMet或Met-tRNAfMet进行蛋白起始。然而，在这些细菌中允许利用Met-tRNAfMet蛋白起始的机制尚不清楚。此外，为什么有些细菌有能力同时合成fMet-tRNAfMet和Met-tRNAfMet，而另一些细菌只合成fMet-tRNAfMet，目前还不清楚。我们研究的主要目标是：(1)理解为什么有些细菌有能力在没有甲酰化的情况下启动蛋白质合成，而另一些细菌则依赖甲酰化；(2)鉴定其他不依赖甲酰化的真细菌来启动蛋白质合成。

The initiator methionyl-tRNA is used to initiate protein synthesis in archaebacteria and the cytoplasm of eukaryotes.  In contrast, protein synthesis in eubacteria and in certain eukaryotic organelles, such as mitochondria and chloroplasts, can be initiated using the initiator formyl-methionyl-tRNA (fMet-tRNAfMet).  Formylation is specific for the initiator methionyl-tRNA (Met-tRNAfMet) and is catalyzed by methionyl-tRNA formyltransferase (MTF). Early studies in vitro and in vivo led to the general belief that formylation of the Met-tRNAfMet is a key step in initiation of protein synthesis in all eubacteria.  This view was reinforced by the findings that disruption of the MTF gene in E. coli, S. pneumoniae and B. subtilis severely curtailed cell growth and the MTF gene is present in all eubacterial genomes sequenced to date.  However, this dogma of eubacterial protein initiation is no longer valid, since some bacteria have been shown to initiate protein synthesis independently of formylation. My laboratory was the first to demonstrate that a eubacteria, Pseudomonas aeruginosa, has the capacity to use Met-tRNAfMet to initiate protein synthesis.  Subsequently, it was demonstrated that formylation of Met-tRNAfMet was not necessary either for protein initiation in S. aureus, H. influenzae and Saccharomyces cerevisiae mitochondria under certain growth conditions.  Thus, unlike E. coli, S. pneumoniae and B. subtilis, which are dependent on fMet-tRNAfMet for protein initiation, P. aeruginosa, S. aureus, and H. influenzae can perform initiation with either fMet-tRNAfMet or Met-tRNAfMet.  The mechanism, however, that allows utilization of Met-tRNAfMet in protein initiation in these bacteria is not understood.  Furthermore, why some bacteria have the ability to initiate protein synthesis with both fMet-tRNAfMet and Met-tRNAfMet and others with only fMet-tRNAfMet is not understood.  The main goals of our research are to (1) understand why some bacteria have the ability to initiate protein synthesis without formylation while others rely on formylation, and (2) identify other eubacteria that are not dependent on formylation for protein initiation.